Diptera: Tephritidae): Low Incidence of Repeat Feeding

Shelly: Methyl eugenol consumption by Bactrocera dorsalis 201

CONSUMPTION OF METHYL EUGENOL BY MALE BACTROCERA DORSALIS (DIPTERA: TEPHRITIDAE): LOW INCIDENCE OF REPEAT FEEDING

TODD E. SHELLY Hawaiian Evolutionary Biology Program University of Hawaii Honolulu, HI 96822

ABSTRACT

The tendency of male Bactrocera dorsalis (Hendel) to re-visit a methyl eugenol source following initial exposure was examined. The first field test investigated the effect of duration of exposure on subsequent capture probability. “Treated” males were allowed to feed on methyl eugenol for 30 s or had access to methyl eugenol for 1 h, 4 h, or 24 h immediately prior to release. Capture probabilities (1%-4%) did not differ significantly among the different treatments but were significantly below that (22%) recorded for “control” (unexposed) males. In a second field test, treated males were released 7 d, 21 d, or 35 d after an initial exposure (2 h) to methyl eugenol. Cap- ture probabilities (11%-18%) did not differ significantly among the different treatments but were significantly below that (34%) recorded for control males. Laboratory tests yielded similar results as both the incidence and duration of re-feeding on methyl eugenol were uniformly low for males held 7 d, 21 d, or 35 d after their initial exposure. By exposing sterile males to the lure prior to release, it may be possible to combine programs of male annihilation and sterile insect release. The present findings also suggest that the effectiveness of male annihilation efforts may be reduced in areas where wild males have consumed sufficient amounts of methyl eugenol from natural sources. Key Words: Oriental fruit fly, parapheromone.

RESUMEN

Se estudió la tendencia de las visitas continuas del macho de Bactrocera dorsalis a una fuente de methyl eugenol. En el primer ensayo se investigó el efecto de el tiempo de exposición en la posibilidad de captura. Los machos tratados fueron ali- mentados con methyl eugenol por 30 segundos y tuvieron acceso a el methyl eugenol por 1, 4 o 24 horas imediatamente antes de la liberacion de los machos. No hubo diferencias significativas para la captura (1-4%) entre los diferentes tratamientos, pero hubo diferencias entre los machos que habian sido expuestos al eugenol com- paradas con los machos sin tratamiento (22%). En el segundo experimento, los machos tratados por dos horas con methyl eugenol fueron liberados a los 7, 21 o 35 días después de tratamiento. No hubo diferencias de captura entre tratamientos, pero la captura fué menor (34%) que en el testigo. Los ensayos de laboratorio dieron resultados simlares en cuanto a la incidencia y la duración de la re-alimentación con methyl eugenol y la captura fué baja para los machos expuestos por 7, 21 o 35 días. Al exponer los machos estériles a el atrayente antes de su liberación, puede ser posible el combinar programas de eliminación de machos y de liberación de machos estériles. Los resultados sugieren que la eficiencia de la eliminación de machos puede aumentar en aquellas areas en las cuales los machos salvajes han consumido can- tidades suficientes de methyl eugenol proveniente de fuentes naturales.

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The males of several economically important tephritid species are strongly attracted to particular chemical compounds, termed “male lures” or “parapheromones”, that either occur naturally in plants or are (presumed) synthetic analogues This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.uﬂ.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

202 Florida Entomologist 77(2) June, 1994 of plant-borne substances (Chambers 1977; Sivinski & Calkins 1986; Fletcher 1987). Several well-known examples include the attraction of male Mediterranean fruit flies, Ceratitis capitata (Wiedemann), to trimedlure, male melon flies, Bactrocera cucurbitae (Coquillett), to cue-lure, and male Oriental fruit flies, B. dorsalis (Hendel), to methyl eugenol. Owing to their powerful attractancy, parapheromones play an important role in current control programs of tephritid pests, both in detecting incipient population outbreaks and eradicating already established populations via male annihilation (Chambers 1977). Despite the wide use of male lures in control efforts, relatively little attention has been given to explaining the underlying biological basis of this sex-specific, chemical attraction. In a recent study on the Oriental fruit fly, Shelly & Dewire (1993) found that “treated” males that fed on methyl eugenol achieved significantly more matings than “control” males deprived of methyl eugenol. Interestingly, treated males had a mating advantage even when they fed on methyl eugenol for only 30 s and were tested 35 d post-feeding. The present study investigates the tendency of B. dorsalis males to re-visit a methyl eugenol source following an initial exposure. Specifically, two field experiments and one laboratory experiment were conducted to examine whether the duration of the initial exposure and the time elapsed since the initial exposure affected the incidence and duration of re-feeding. Based on the results of mating trials (Shelly & Dewire 1993), I predicted that neither the duration of the initial exposure (at least for exposure periods exceeding 30 s) nor the time elapsed since the initial feeding (at least for intervals up to 35 d) would significantly affect the tendency for re-feeding.

MATERIALS AND METHODS

Field Experiments

All flies used in field tests were from a colony maintained by the USDA/ARS Trop- ical Fruit and Vegetable Laboratory, Honolulu, for approximately 70 generations (M. Fujimoto, pers. comm.) using standard rearing procedures (Tanaka et al. 1969). Non- irradiated pupae were obtained 2 d prior to eclosion, and adults were sexed within 5 d of eclosion [(sexual maturity in this stock is attained at about 10 d of age, (M. Fujimoto, pers. comm.)]. Males were kept in 5-liter plastic buckets (50 per bucket) covered with screen mesh and given food and water ad libitum. Experiments were conducted at 2 locations on the island of Oahu, Hawaii. Dur- ing September-October, 1991, I used a 0.6-ha citrus grove in the University of Hawaii Agricultural Experiment Station, Waimanalo, that contained approximately 60 orange trees (Citrus sinensis (L.)). The grove was bordered on two sides by an open field containing small patches of guava (Psidium guajava L.) and coffee (Coffea arabica L.) and on the other two sides by highly disturbed, second-growth forest. During May-July, 1992, field-work was conducted at the Kanewai Garden near the campus of the University of Hawaii, Honolulu. This small area (0.4 ha) contained six large mango trees (Mangifera indica L.) and was bordered by an open lot on one side and lawns containing non-host vegetation on the remaining sides. Two field experiments were performed. At Waimanalo, I examined whether the duration of exposure to methyl eugenol affected capture probability. As described below, treated males fed on methyl eugenol for only 30 s or had access to methyl eugenol for 1 h, 4 h, or 24 h immediately prior to release. At the Kanewai Garden, I examined the effect of time lapse following initial feeding on capture probability. Treated males had access to methyl eugenol for 2 h and were released 7 d, 21 d, or 35 d later. An additional set of treated males was permitted to feed on methyl eugenol for only 30 s and was released 35 d later. Control males that had no exposure to methyl eugenol were also released in both experiments.

Shelly: Methyl eugenol consumption by Bactrocera dorsalis 203

To obtain treated males, 1.5 ml of methyl eugenol was applied to 5-cm long cotton wicks, and the wicks, held upright in small plastic containers, were placed singly in the appropriate buckets during midday. Buckets were placed on a shaded outdoor porch where air temperatures varied between 29-31 oC (or 23-31 oC during 24 h exposure periods). The feeding activity of individual males was not monitored during exposure periods of 1 h or more. To obtain males with 30 s feeding times, groups of 5- 10 males were observed in screen cages (30 cm cubes with a cloth sleeve on one side) containing a single wick. Individuals were removed after 30 s of feeding by gently “coaxing” them into a vial. In all cases, treated males were exposed to methyl eugenol at 14 d of age and correspondingly were released at the age of 14 d at the Waimanalo site and 21 d, 35 d, or 49 d at Kanewai Garden. At Waimanalo, control males were 14 d old at release, while at Kanewai Garden separate control groups of males aged 21 d, 35 d, and 49 d, respectively, were used for the two treatment categories. Prior to release, control males and the males in the different treatment groups were cooled and marked on the thorax with different color combinations of enamel paint (a given combination was used only once at either field site). The cooling and painting procedures had no apparent adverse effects on male behavior, and individuals resumed “normal” activities within minutes of handling. The following protocol was used for the tests conducted at Waimanalo. On the day prior to a release, Steiner traps were placed singly in 16 different trees located throughout the grove. The same trees were used in all tests. Traps were suspended in the canopy by a 30-cm long wire fastened to a branch. Each trap contained a 5-cm long cotton wick to which 1.5 ml of methyl eugenol (3% naled) had been applied. For all tests, the males were released beneath a centrally located orange tree between 1500-1700 hours. The actual release was accomplished by removing the screen top and gently tapping the bucket to induce flight. Males that were unable to fly were not counted in the release sample. Traps were checked 5 d after release, and in the laboratory captured flies were examined individually for markings. Six replicates were conducted with 75-112 males released per group (control or treatment) per replicate. A similar release protocol was employed at the Kanewai Garden site. However, owing to the small size of the garden, only eight Steiner traps were used at this site. The traps were placed in a circle (70-m radius) around a central release point (a mango tree). Eight replicates were conducted for tests involving treated males exposed to methyl eugenol for 2 h and released 7 d or 21 d later, with 122-143 males released per group (control or treatment) per replicate. Four replicates were conducted for tests involving treated males released 35 d after either exposure to methyl eugenol for 2 h (82-113 males per group per replicate) or feeding on methyl eugenol for only 30 s (79-120 males released per group per replicate).

Laboratory Observations

The effects of feeding duration and time since ﬁrst feeding on the incidence and duration of repeat feedin.g were also investigated in the laboratory. Males used in these tests were from a laboratory stock started in November, 1991, with 200-300 adults reared from mangos collected in Waimanalo. Data were collected in July-Sep- tember, 1992; consequently, the individuals observed were approximately eight generations removed from the wild. Larvae were reared on papaya, and adults were separated by sex within 7 d of eclosion, well before reaching sexual maturity (at approximately 15-20 d of age, Foote & Carey 1987). Treated males fed on methyl eugenol for only 30 s (following the protocol described above) or had access to methyl eugenol for a 30-min period during which their feeding activity was monitored. To obtain this latter group, ﬁve uniquely marked individuals were placed in screen cages (30-cm cubes), allowed a 1-2 h “accli- mation period”, and then given free access to a 5-cm long cotton wick to which 1.5 ml of methyl eugenol had been applied. The amount of time that individual males fed on

204 Florida Entomologist 77(2) June, 1994 the wick was then recorded to the nearest second. All observations were made between 1100-1330 hours on a shaded outdoor porch at temperatures between 29-31 oC. Following the initial exposure, treated males were kept in 5-liter plastic buckets and given ample food and water. Treated males - both those restricted to 30 s feeding and those given 30 min access - were held 7 d, 21 d, or 35 d before a second exposure (30 min) during which feeding times of individual males were recorded. All treated males were initially exposed to methyl eugenol at 25 d of age. To investigate the possibility that male age was partly responsible for any feeding differences observed between the ﬁrst and second exposures, I recorded the feeding times of uniquely marked, control males given their ﬁrst exposure (30 min) to methyl eugenol at ages 32 d, 46 d, and 60 d, respectively (ages correspond to those of treated males held 7 d, 21 d, or 35 d, respectively).

RESULTS

Field Experiments

In the Waimanalo experiment, no signiﬁcant differences in capture probability were found among males in the different treatment groups (H=6.1; P > 0.05; Kruskal-Wallis test; Fig. 1). Among the different exposure groups, only 1%-4% of the males were captured, on average, in a given replicate. In contrast, 22% of control

Fig. 1. Capture probabilities of B. dorsalis males exposed to methyl eugenol for varying lengths of time. Points represent average proportion of males captured per replicate; vertical lines indicate + standard error. Release groups: C=control, T=treated. T-30 s males were restricted to 30 s of feeding on methyl eugenol; the remaining groups of treated males had access to methyl eugenol for 1 h, 4 h, or 24 h, respectively. See text for sample sizes.

Shelly: Methyl eugenol consumption by Bactrocera dorsalis 205 males were captured, on average, in a given replicate. The capture probability of control males differed significantly from males exposed for 1 h (q=5.6), 4 h (q=6.0), or 24 h (q=5.2) as well as from males whose feeding was restricted to 30 s (q=7.9; P < 0.005 in all cases; multiple comparisons test, Zar 1974: 156). At the Kanewai Garden, no significant differences in capture probability were detected among males exposed to methyl eugenol for 2 h but released after differing time intervals (H=5.1; P > 0.05; Kruskal-Wallis test; Fig. 2). Over the different intervals, only 11%-18% of the treated males were captured, on average, in a given replicate. Similarly, capture probabilities did not differ among control males held for varying periods before release (H=0.5; P > 0.05; Kruskal-Wallis test; Fig. 2). On average, approximately 33% of control males were trapped over all pre-release intervals. Based on data pooled over all pre-release intervals, the capture probability for control males was significantly higher than that observed for males given 2 h access to methyl eugenol before release (U=387.5; P < 0.001; Mann-Whitney test). Treated males that fed for only 30 s prior to their release 35 d later also had low capture probability (Fig. 2). An average of 11% of these males was captured per replicate, the same proportion observed for males released 35 d after 2 h exposure to methyl eugenol (U=9; P > 0.05; Mann-Whitney test).

Laboratory Observations

Among treated males given an initial 30-min exposure period, feeding durations were signiﬁcantly shorter during the second exposure for males tested 7 d (T=276; n=54), 21 d (T=87; n=53), or 35 d (T=3; n=27) after the initial feeding (P < 0.001 in all

Fig. 2. Capture probabilities of B. dorsalis males held varying lengths of time after exposure to methyl eugenol. Points represent average proportion of males captured per replicate; vertical lines indicate + standard error. One set of treated males (held 35 d) was restricted to 30 s of feeding on methyl eugenol; all other treated males had access to methyl eugenol for 2 h. See text for sample sizes.

206 Florida Entomologist 77(2) June, 1994 cases; Wilcox on paired-sample test; Fig. 3). Moreover, for these males, feeding durations during the second exposure were independent of time elapsed since the initial feeding (H=1.1; P > 0.05; Kruskal-Wallis test). Among the different trials, 85%-91% of the males consumed methyl eugenol during the initial exposure compared to only 32%-38% during the second exposure. Decreased feeding during the second exposure was apparently not age-related: average feeding durations were similar among control males aged 32 d (n=35), 46 d (n=40), and 60 d (n=40; H=3.9; P > 0.05; Kruskal- Wallis test; Fig. 3). Data pooled over the different inter-exposure intervals (or, equiv- alently, male ages) revealed that, during their second exposure period, treated males fed for shorter periods of time, on average, than control males (Z=11.1; P < 0.05; n1=134, n2=115; Mann-Whitney U-test). Among treated males given an initial 30-min exposure, there was no correlation in the feeding times of individual males between the first and second exposure periods (rs=0.05; P > 0.05; n=134; Spearman rank). Even if only the incidence of feeding is considered (i.e., regardless of duration), feeding activity during the first exposure period was still not a reliable predictor of subsequent feeding activity: males that fed during the first exposure period were as likely to feed during the second period (48 of 118=41%) as were males that did not feed at all during the initial exposure (8 of 16=50%; G=0.4; P > 0.05; G test with Yates correction). Among treated males given two 30-min exposure periods, 6% (8/134) did not feed on methyl eugenol during either period.

Fig. 3. Feeding times of B. dorsalis males during their second exposure to methyl eugenol 7 d, 21 d, or 35 d after the initial exposure. One set of treated males was given an initial 30 min exposure period, while another set was restricted to an initial feeding of 30 s; for both sets of treated males, the second exposure period was 30 min. Data for control males represent feeding durations during initial 30-min exposure periods at ages corresponding to males in different treatment groups. Points represent average values; vertical lines indicate + standard error. The value plotted for the initial exposure was calculated over all treated males given an initial 30-min exposure period. See text for sample sizes.

Shelly: Methyl eugenol consumption by Bactrocera dorsalis 207

Treated males limited to an initial feeding of 30 s also displayed low feeding activity during the second exposure period (Fig. 3). In fact, when re-exposed to methyl eugenol 7 d (n=35 males) or 21 d (n=35 males) after the first feeding, these individuals had feeding durations that were similar to (and not greater than, as might be expected) males given an initial access of 30 min (7 d - Z=0.6; n1=35, n2=54; 21 d - Z=0.5; n1=35, n2=53; P > 0.05 in both cases; Mann-Whitney U-test). However, at 35 d after the initial exposure, males (n=40) limited initially to a 30 s feeding fed longer, on average, than males first given a 30 min exposure period (Z=2.7; P < 0.01; n1=40, n2=27; Mann-Whitney U-test). Though feeding durations of these males increased after 35 d, they were still significantly lower than those of control males of the same age (Z=2.1; P < 0.05; n1=n2=40; Mann-Whitney U-test).

DISCUSSION

Results of the present study indicate that after an initial exposure, B. dorsalis males have a greatly reduced tendency to re-visit a methyl eugenol source. In the field experiments, males that were permitted only 30 s feeding on methyl eugenol were rarely captured in methyl eugenol-baited traps even when released 35 d after feeding. Similarly, in the laboratory most males given an initial exposure of 30 min “ignored” a methyl eugenol source placed directly in their cage 35 d later. Though data are scant, it appears that a dramatic reduction in male responsiveness to lures following exposure characterizes other tephritid species as well. Us- ing a large outdoor cage, Chambers et al. (1972) reported that, after initial exposure to cue-lure, only 14% of male B. cucurbitae, on average, responded to cue-lure-baited traps compared to 50% of control (unexposed) males. Similarly, Brieze-Stegeman et al. (1978) placed dye in a methyl eugenol-baited trap (lacking poison) and found that only 13% (daily average) of the B. cacuminatus (Hering) males seen at the trap over the next several days were marked. The major difficulty in interpreting laboratory studies on male attraction to lures is the scarcity of field data regarding both the availability of parapheromones in natural sources and the feeding behavior of males at these sources. To my knowledge, no data exist regarding either the incidence and duration of feeding bouts or the rate and amount of parapheromone consumption during these bouts. It is likely that the 1-2 ml doses of parapheromones used by experimenters (Chambers et al. 1972; Brieze-Stegeman et al. 1978; present study) exceed levels available in natural sources (e.g., Kawano et al. 1968). Despite this possible discrepancy, it is certainly conceivable that in the wild, males initially make frequent or prolonged feeding bouts and in so doing eventually consume parapheromone in amounts similar to males observed in laboratory studies. In other words, though the feeding time required to inhibit subsequent feeding is reduced in laboratory studies, the basic pattern of decreased responsiveness to parapheromones may nonetheless be characteristic of wild males. The present study has three major implications for control or eradication projects of tephritid pests. First, by exposing sterile males to the lure prior to their release, workers may be able to combine programs of male annihilation and sterile insect release. As noted by Chambers et al. (1972), pre-exposure of sterile males may increase the efficiency of achieving effective overflooding ratios, since wild males would respond to lure-baited traps, whereas sterile males would not. Pre-exposure to the parapheromone may also increase the mating competitiveness of sterile males (Shelly & Dewire 1993), further enhancing the effectiveness of the sterile insect release method. Second, the present findings suggest the possibility that wild males that have consumed sufficient amounts of parapheromone from natural sources may show reduced attraction to lure-baited traps, thus potentially reducing the effectiveness of male annihilation programs. Finally, and somewhat unexpectedly, 6% of the

208 Florida Entomologist 77(2) June, 1994 males observed in the laboratory tests were not attracted to methyl eugenol in two separate exposure periods. The possibility that some males in a population may respond only slightly or not at all to parapheromones implies that in certain situations male annihilation may fail to achieve total eradication. Studies in our laboratory are currently investigating the genetic basis of male responsiveness to parapheromones using the B. dorsalis-methyl eugenol association.

ACKNOWLEDGMENTS

I thank the staff of the University of Hawaii Agricultural Experiment Station in Waimanalo for their cooperation. Annie Dewire, Stacey Fong, Caryn Ihori, Cheryl Monez, and Michael Whang provided capable laboratory assistance, and to all I am grateful. Also, I thank Emma Shelly who, despite her young age, was a great help in counting marked ﬂies in trap catches. Comments by Tim Whittier greatly improved the paper. This research was supported by funds from the California Department of Food and Agriculture (90-0581) and the USDA/ARS (58-91H2-6-42).

REFERENCES CITED

BRIEZE-STEGEMAN, R., M. J. RICE, AND G. H. S. HOOPER. 1978. Daily periodicity in attraction of male tephritid fruit flies to synthetic chemical lures. J. Austra- lian Entomol. Soc. 17: 341-346. CHAMBERS, D. L., K. OHINATA, M. FUJIMOTO, AND S. KASHIWAI. 1972. Treating tephritids with attractants to enhance their effectiveness in sterile-release programs. J. Econ. Entomol. 65: 279-282. CHAMBERS, D. L. 1977. Attractants for fruit fly survey and control, pp. 327-344 in H. H. Shorey and J. J. McKelvey [eds.], Chemical control of insect behavior. Wiley, New York. FLETCHER, B. S. 1987. The biology of dacine fruit flies. Ann. Rev. Entomol. 32: 115- 144. FOOTE, D., AND J. R. CAREY. 1987. Comparative demography of a laboratory and a wild strain of the Oriental fruit fly, Dacus dorsalis. Entomol. Exp. Appl. 44: 263-268. KAWANO, Y., W. C. MITCHELL, AND H. MATSUMOTO. 1968. Identification of the male Oriental fruit fly attractant in the golden shower blossom. J. Econ. Entomol. 61: 986-988. SHELLY, T. E., AND A. DEWIRE. 1993. Chemically mediated mating success in male Oriental fruit flies, Bactrocera dorsalis (Diptera: Tephritidae). Ann. Entomol. Soc. America. In press. SIVINSKI, J. M., AND C. CALKINS. 1986. Pheromones and parapheromones in the control of tephritids. Florida Entomol. 69: 157-168. TANAKA, N., L. F. STEINER, K. OHINATA, AND R. OKAMOTO. 1969. Low-cost larval rearing medium for mass production of Oriental and Mediterranean fruit flies. J. Econ. Entomol. 62: 967-968. Zar, J. H. 1974. Biostatistical analysis. Prentice-Hall, Inc. Englewood Cliffs, NJ.

Fontes et al.: Phytophagous Insects Associated with Goldenrods209

PHYTOPHAGOUS INSECTS ASSOCIATED WITH GOLDENRODS (SOLIDAGO SPP.) IN GAINESVILLE, FLORIDA

E. M. G. FONTES1, D. H. HABECK, AND F. SLANSKY, JR. Dept. of Entomology & Nematology University of Florida Gainesville, FL 32611-0740

ABSTRACT

The insect fauna of four species of goldenrods, Solidago canadensis var. scabra, S. ﬁstulosa, S. gigantea and S. leavenworthii, was surveyed during four years in and around Gainesville, Florida. The 122 phytophagous species collected are listed and classiﬁed according to relative frequency of occurrence, guild, host range, plant part attacked, life stages collected, and associated goldenrod species. Only 14 (11%) of the phytophagous species are known to be restricted to goldenrods and Aster (Composi- tae). Eight insect species are considered as possible biological control agents of Sol- idago spp.

RESUMEN

La fauna de insectos presente en cuatro especies de vara de oro, Solidago canadensis var scabra, S. fistulosa, S. gigantea y S. leavenworthii fué, estudiada en Gainesville, Florida durante cuatro años. Los 122 specimenes fitófagos colectados, se han listado y clasificado de acuerdo a la frequencia relativa de aparición, asociación, rango de hospedantes, parte de la planta atacada, estado de desarrollo y especies de vara de oro a las que se asociaron. Solamente 14 (11%) de los fitófagos hallados son conocidos como específicos de las vara de oro y Aster (Compositae). Ocho especies son consideradas como posibles agentes de control biológico de Solidago spp.

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Goldenrods (Asteraceae: Solidago spp.) are common on roadsides and in open fields throughout the eastern United States. They first attracted the attention of nat- uralists because of their aesthetic appeal and as a nectar source for pollinators in late fall (Feller-Demalsy & Lamontagne 1979, Hensel 1982). Goldenrods have been studied as sources of natural enemies of field crop pests in a mixed crop system (Alt- ieri 1979) and as competitors with seedlings of woody trees (Norbi & Kozlowski 1980). They can also serve as a reservoir for disease-producing organisms that attack plants of economic importance (Werner et al. 1980). Introduced to Europe and Japan about 1900, goldenrods have become aggressive pests of forest nurseries and refores- tation areas (Capek 1971). The high cost and low efficiency of chemical and mechan- ical control of these weeds have resulted in interest in a biological control program in Russia (O. Kovalev, pers. comm.). Basic information on the arthropod community structure of goldenrods is relevant if goldenrod populations are to be manipulated either as beneficial plants that should be maintained or even enhanced, or as undesirable plants that should be controlled. This paper lists the phytophagous insect fauna of four species of goldenrods, (Solidago canadensis L. var. scabra (Muhl) [treated by many as S. altissima L.], S. fistulosa Miller, S. gigantea Aiton, and S. leavenworthii Torr. & Gray), in and around Gainesville, Florida. It also gives information on the relative abundance of the different taxa, parts of the plant attacked, and specificity of the phytophagous insects.

1Present address: EMBRAPA, CENARGEN, S.A.I.N. Parque Rural, C.P. 10.2372 Brasilia - DF - CEP 70.770, BRAZIL. This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.uﬂ.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

210 Florida Entomologist 77(2) June, 1994

MATERIAL AND METHODS

The insect fauna of goldenrods was surveyed for four consecutive growing seasons. Surveys began in June 1981 and were made on goldenrod populations as they occurred naturally. Plants, including roots, were checked in the field and/or placed in plastic bags and returned to the laboratory for examination. Insect-damaged plant parts were carefully examined under the microscope. Immature insects were reared individually, with information recorded as to the life stage collected, part of the plant damaged, life history, appearance, emergence of adults or parasites, presence of disease, and other relevant information. Flying insects were captured by making 10 sweeps with a sweep net while walking in a straight line through the goldenrod patch. Adults were preserved in alcohol and identified. Plants were sampled at different times of the year and at various growth stages (seedling, flowering and mature). In 1982, 16 populations of the four plant species were selected of which two were surveyed each week following the same procedure. Of the four goldenrod species, S. canadensis var. scabra was the most common (11 of the 16 populations) and occurred on ditch banks, roadsides, and clearings in woody areas. The second most common species, S. fistulosa, occurred in old abandoned fields forming large stands sometimes mixed with blackberries. The single stand of S. gigantea was small and located on a poorly drained roadside. The S. leavenworthii stand was mixed with many other weeds and was growing on a small ditch bank in a very disturbed area. Growth of all four species started in March; S. leavenworthii and S. gigantea flow- ered in late August, S. fistulosa in mid-September and S. canadensis var. scabra in late September. The goldenrod stems were completely dead by the beginning of De- cember. In 1983, three populations of S. canadensis, two of S. fistulosa, and single populations of S. gigantea and S. leavenworthii, were selected for in-depth study. The dynamics of the insect fauna in relation to host plant phenology was followed by sur- veying these seven populations during four periods of the growing season: early (May), middle (July), about 10 days before blooming (August or early September), and shortly after blooming (late September or early October). Collections in 1984 were made only on S. canadensis var. scabra and S. leavenworthii. A phytophagous species was considered common if it was present in at least 50% of the collections, occasional if collected in 15 to 50% of the samples, and rare if present in less than 15% of the samples. Insects collected only once during the four years were excluded, as were those collected often but known to be nectar/pollen feeders or incidental and associated with other plants.

RESULTS AND DISCUSSION

Seven orders, encompassing 41 families with 123 species of phytophagous insects (other than nectar/pollen feeders) were found associated with Solidago spp. (Table 1). Of these, 60 species (49.2%) bred on these plants, as indicated by the presence of immature stages. The cercopid Lepyronia quadrangularis (Say), the cicadellid Os- bornellus clarus Beamer, and the delphacid Pissonotus marginatus Van Duzee were new Alachua County records, and the gracillariid Cremastobombycia solidaginis (Frey and Boll.) was a new state record. Of the phytophagous insect species, 16 (13.1%) fed only on Solidago or on Sol- idago and Aster, seven (5.7%) fed on these plants as well as other Compositae, 94 (77%) were polyphagous and 5 (4.1%) had unknown hosts. Sucking insects comprised the majority of the phytophagous insects with 35 species of Homoptera and 25 species of Hemiptera collected from leaves, stems and ﬂowers. There were 14 species of cicadellids, six of which were common. The pentatomids and mirids were represented by six species each, and the lygaeids by ﬁve.

Fontes et al.: Phytophagous Insects Associated with Goldenrods211

TABLE 1. PHYTOPHAGOUS INSECT FAUNA OF SOLIDAGO SPP. IN THE VICINITY OF GAINESVILLE, FL

Relative Stages Plant Solidago Host Freq.1 Coll.2 Part(s)3 Guild4 spp.5 Spec.6

DIPTERA

Agromyzidae Agromyzidae sp. 1 C L,P,A L M c,f,g,l M Agromyzidae sp. 2 C L,P,A L M c,f,g,l -

Cecidomyiidae Asphondylia monacha C L,P,A L G c M Osten Sacken Asteromyia carbonifera C L,P,A L G c,f,g,l M (Osten Sacken) Asteromyia sp. nr. carbonifera (Osten Sacken) C L L G c,f,g,l M Dasyneura sp. C L L G c,f,g,l M Rhopalomyia sp. (prob. new) O L,P,A St G f M Rhopalomyia solidaginis (Loew) O L,P,A St G f,g,l M Schizomyia racemicola (Osten Sacken) O L,A F G c,f M

Tephritidae Eurosta prob. comma (Wied.) R L R G f M

COLEOPTERA

Cerambycidae Strangalia sexnotata Hald R A F C f P

Chrysomelidae Arthrochlamys plicatus (Fab.) R L L C c,f P Colaspis brunnea (Fab.) R A L C f P Colaspis favosa (Say) R A L C c P Diabrotica undecimpunctata howardi Barber R A L C c,f P Diachus auratus Fab. O A L C c,f,l P Exema canadensis Pierce C L,P,A L C c,f,l P 212 Florida Entomologist 77(2) June, 1994

TABLE 1. (CONTINUED)PHYTOPHAGOUS INSECT FAUNA OF SOLIDAGO SPP. IN THE VICINITY OF GAINESVILLE, FL

Relative Stages Plant Solidago Host Freq.1 Coll.2 Part(s)3 Guild4 spp.5 Spec.6 Exema sp. R L L C c,l F Ophraella sexvittata (LeC.) C E,L,A L C c,f,g,l M Paria sp. nr. aterrima (Oliver) O A L C c,f P Rhabdopterus sp. R A L C c P Systena elongata (Fab.) R A L C c,f A

Curculionidae Centrinaspis picumnus (Herbst) C A L,F C c,f,l P Epicaerus formidolosus Boheman R A L C c,f P Limnobaris sp. O A L,F C c,f,g P Notolomus basalis LeC. C A F C c,f,g P Pachnaeus opalus (Oliver) R A L C c P Tanymecus “lacaena (Hbst.)” complex O A L,F C c P

Elateridae Conoderus lividus (De Geer) R A L C f,l P Glyphonyx sp. R A L C c P

Meloidae Epicauta sp. O A F C c,f P

Mordellidae Mordellistena sp. R A L C c,l -

Phalacridae Olibrus sp. R A L C c,f -

Scarabaeidae Trigonopeltastes delta (Forst.) O A F C c,f,l P

HEMIPTERA

Alydidae Alydus pilosulus (Herrich-Shaffer) O A L S c,g,l P Fontes et al.: Phytophagous Insects Associated with Goldenrods213

TABLE 1. (CONTINUED)PHYTOPHAGOUS INSECT FAUNA OF SOLIDAGO SPP. IN THE VICINITY OF GAINESVILLE, FL

Relative Stages Plant Solidago Host Freq.1 Coll.2 Part(s)3 Guild4 spp.5 Spec.6 Leptocorisa ﬁliformis (Fab.) R L,A F S f P

Coreidae Acanthocephala femorata (Fab.) O E,L,A L S c,f,l P Leptoglossus phyllopus (L.) C A L S c,f,g,l P

Lygaeidae Cymonimus notabilis (Distant) R A L S c,f P Ochrostomus lineoloides Slater R A F,L S f,l P Ocrimnus mimulus (Stal) C L,A L,S S c,f,l Oedancala crassimana (Fab.) R L,A L S f P Pachybrachius bilotatus (Say) C L,A F,L S c,g,l P

Miridae Adelphocoris rapidus (Say) O A L S c,f,l P Lepidopsallus pusillus (Knight) O A L S c,f,l P Lygus lineolaris (Palisot de Beauvois) C A L S f,g,l P Polymerus punctipes Knight R A L S f P Rhinacloa pusillia (Knight) R A F S c,l P Taylorilygus pallidulus (Blanchard) C L,A L S c,f,g,l P

Pentatomidae Euschistus obscurus (Palisot de Beauvois) R A L S c,l P Euschistus servus (Say) C L,A L S c,f,g P Holcostethus limbolarius (Stal) R A L,F S f,l P Nezara viridula (L.) C E,L,A L S c,g,l P Oebalus pugnax (F.) R L,A L S f P Thyanta custator (F.) C L,A L S c,g,l P 214 Florida Entomologist 77(2) June, 1994

TABLE 1. (CONTINUED)PHYTOPHAGOUS INSECT FAUNA OF SOLIDAGO SPP. IN THE VICINITY OF GAINESVILLE, FL

Relative Stages Plant Solidago Host Freq.1 Coll.2 Part(s)3 Guild4 spp.5 Spec.6 Tingidae Corythucha marmorata (Uhler) C E,L,A L S c,f A

Rhopalidae Arhyssus lateralis (Say) R A F S c,f - Arhyssus nigristernum (Signoret) O A F S g,l P Harmostes reﬂexulus (Say) R A F S c,f P

HOMOPTERA

Aphididae Aphis sp. O L,A L S c,f P Hyperomyzus lactucae (L.) C L,A St S c,f,l P Uroleucon ambrosiae (C. Thomas) C L,A St S c,f,g,l P Uroleucon gravicornis (Patch) C L,A St S c,f,g,l P

Cercopidae Clastoptera xanthocephala German C L,A St S c,f,g,l P Lepyronia quadrangularis (Say) C L,A St S c,f,l P

Cicadellidae Agallia constricta Van Duzee R A L S g,f P Chlorotettix viridius Van Duzee R A L S f P Empoasca sp. C L,A L S c,f,g,l P Exitianus exitiosus (Uhler) R A L S l P Graminella sonorus (Ball) R A F S l P Graphocephala versuta (Say) C A L S c,f,g,l P Gyponana sp. C L,A L S c,f,g,l P Fontes et al.: Phytophagous Insects Associated with Goldenrods215

TABLE 1. (CONTINUED)PHYTOPHAGOUS INSECT FAUNA OF SOLIDAGO SPP. IN THE VICINITY OF GAINESVILLE, FL

Relative Stages Plant Solidago Host Freq.1 Coll.2 Part(s)3 Guild4 spp.5 Spec.6 Homalodisca coagulata (Say) C A L S c,f,g,l P Homalodisca insolita (Walker) R A L S c P Oncometopia nigricans (Walker) C E,L,A L S c,f,g,l P Osbornellus clarus Beamer O A L S c,f,g P Scaphytopius sp. nr. acutus (Say) C L,A L S c,f,g P Scaphytopius sp. prob. frontalis (Van Duzee) O A L S c,l P Sibovia occatoria (Say) R A L S f,l P

Coccidae Coccus hesperidum L.RA L S g P

Delphacidae Pissonotus marginatus Van Duzee R A L S c,f P Sogatella kolophon meridiana (Beamer) R A L S c P Diaspididae Abgrallaspis cyanophylli (Signoret) R A L S f P Aonidomytilus solidaginis (Hoke) O L,A St S c,f A Hemiberlesia lataniae (Signoret) R A St S f P Pseudaulacaspis pentagona (Targ. Tozz.) R A L S f P

Dictyopharidae Rhynchomitra sp. C L,A L S c,f,g P

Flatidae Ormenoides venusta (Melichar) R A L S c P

Membracidae Acutalis tartarea (Say) C L,A St S c,f,g,l P Entylia bactriana Germar O L,A St S c,f P 216 Florida Entomologist 77(2) June, 1994

TABLE 1. (CONTINUED)PHYTOPHAGOUS INSECT FAUNA OF SOLIDAGO SPP. IN THE VICINITY OF GAINESVILLE, FL

Relative Stages Plant Solidago Host Freq.1 Coll.2 Part(s)3 Guild4 spp.5 Spec.6 Micrutalis calva (Say) C A L S c,f,g,l P Spissistilus festinus (Say) O A L S c,f P

Psyllidae Aphalara sp. O L,A St S f,l P Craspedolepta veaziei (Patch) O A L S f,g M

LEPIDOPTERA

Gelechiidae Gnorimoschema gallaesolidaginis (Riley) O L,P,A St G f,g M Trichotaphe ﬂavocostella (Clem.) C L,P,A L L c,f A Trichotaphe inserrata (Wlsm.) C L,P,A St B c A

Geometridae Eupithecia miserulata Grote O L,P,A F C c,f P Pleuroprucha insulsaria (Guenee) R L,A F C f,g P Synchlora frondaria (Walk.) O L,P,A L,F C c,f,l P

Gracillariidae Cremastobombycia solidaginis (Frey and Boll.) C L,P,A L M c,f,g M

Lyonetiidae Bucculatrix solidaginiella Braun R P,A L M c M

Noctuidae Schinia nundina (Drury) R L,P,A F,Sd C f M

Pterophoridae Oidaematophorus kellicottii (Fish) C L,A St B c,f,g,l A Fontes et al.: Phytophagous Insects Associated with Goldenrods217

TABLE 1. (CONTINUED)PHYTOPHAGOUS INSECT FAUNA OF SOLIDAGO SPP. IN THE VICINITY OF GAINESVILLE, FL

Relative Stages Plant Solidago Host Freq.1 Coll.2 Part(s)3 Guild4 spp.5 Spec.6

Tortricidae Platynota ﬂavedana Clem. R L L C c P Platynota rostrana (Walk.) R L,P F C c P Sparganothis distincta (Walsing.) C L,P,A L L c,f,g,l M

ORTHOPTERA

Acrididae Aptenopedes sphenarioides Scudder C L,A L C c,f,g P Melanoplus sp. C L L C c,f,g,l P Paroxya atlantica Scudder C L,A L C f,g,l P Schistocerca damniﬁca (Saussure) R A L C f P

Blatellidae Cariblatta lutea (Saussure Zehntner) O L,A L C c P

Grylliidae Hapithus brevipennis (Saussure) C L,A L C c,f,g,l P Oecanthus celerinictus Walker O L,A L C c,f P Orocharis luteolira Walker R A L C c P

Tettigoniidae Amblycorypha prob. ﬂoridana Rehn Hebard R L,A L C c P Belocephalus subapterus Scudder O L L C c,g P Odontoxiphidium apterum Morse O A L C c,f P Scudderia sp. O A L C c,f P

THYSANOPTERA 218 Florida Entomologist 77(2) June, 1994

TABLE 1. (CONTINUED)PHYTOPHAGOUS INSECT FAUNA OF SOLIDAGO SPP. IN THE VICINITY OF GAINESVILLE, FL

Relative Stages Plant Solidago Host Freq.1 Coll.2 Part(s)3 Guild4 spp.5 Spec.6 Phlaeothripidae Elaphrothrips sp. R A L S l -

Thripidae Microcephalothrips abdominalis (Crawford) R A L S l A Thrips tabaci Lindeman R A L S c P

1C = common; O = occasional; R = rare. 2E = eggs; L = larvae; P = pupa; A = adult. 3L = leaves; St = stem; F = ﬂowers; R = roots; Sd = seeds. 4C = chewing; G = gallmaker; M = miner; B = borer; S = sucking; L = leaftier. 5c = Solidago canadensis var. scabra; f = S. ﬁstulosa; g = S. gigantea; l = S. leavenworthii. 6M = monophagous: feed only on Solidago and Aster; A = feed on Solidago and on other genera of Asteraceae; P = Polyphagous: feed also on families other than Asteraceae.

Two mirids and three pentatomids are associated with economic damage to cultivated plants. Lygus lineolaris Palisot de Beauvois, the tarnished plant bug, causes serious damage by feeding on the tender growing or fruiting parts of a variety of wild and cultivated plants (Borror et al. 1976, Metcalf & Flint 1951). Adults of this insect were collected from May to October on S. fistulosa, S. gigantea, and S. canadensis var. scabra. Taylorilygus pallidulus (Blanchard) attacks a variety of wild plants and, when abundant, can be a pest of ornamentals (F. W. Mead, pers. comm.). It fed and bred on all four species of goldenrod and was very common throughout the growing season. Among the pentatomids, the southern green stink bug, Nezara viridula (L.), is the most important because it is a pest of soybeans and other crops (Todd & Herzog 1980). It fed and bred from May to July on three of the four species of goldenrod. Eu- schistus servus (Say) and Oebalus pugnax (Fab.) also can cause damage to many crops. Most of the chewing insects were polyphagous. Of 12 species of chrysomelids, apparently only Ophraella sexvittata (LeConte) was restricted to Solidago, although LeSage (1986) reported adults were reared from larvae on march elder, Iva frutescens L. sp. oraria (Bart.) R. C. Jackson (Asteraceae). Systena elongata (Fab.) was rare and collected only on S. canadensis var. scabra and S. leavenworthii. It feeds only on plants in the Asteraceae. Diabrotica undecimpunctata howardi Barber, the spotted cucumber beetle, was the only economic species of chrysomelid collected on Solidago. Adults were collected several times on S. canadensis var. scabra and S. fistulosa. Polyphagous Lepidoptera collected were restricted to three geometrid and two tortri- cid species. All species of Orthoptera collected were polyphagous. The most abundant insect feeding only on Solidago and other Asteraceae was the tingid Corythucha marmorata (Uhler). Its' eggs, nymphs and adults were found on S. canadensis var scabra and S. fistulosa throughout the growing season. The pterophorid Oidaematophorus kellicottii (Fish) was common on all four species. This moth laid its' eggs on the growing tip of young goldenrod plants. The new larvae bored downward into the soft stem. Before reaching the apparent third instar, it left the upper part of the plant stem through a lateral hole and moved down to the mature, wider stem. Here it made another hole, about 10 cm above the ground, and bored downward toward the roots. Pu- pation occurred inside the stem near this entrance hole, through which the adult emerged later. The part of the plant above where the young larva bored, wilted and Fontes et al.: Phytophagous Insects Associated with Goldenrods219 died. Infested plants could be recognized by the dried tips. Young larvae were collected in May and mature ones were found as late as October. Other common insects were the gelechiids Trichotaphe flavocostella (Clem.) and T. inserrata (Wlsm.). These species caused considerable damage to goldenrods; the former as a leaftier and the latter as a borer inside the growing tip, a behavior that interrupted the terminal growth of the plant. Most of the species restricted to Sol- idago were endophagous: eight Diptera and one Lepidoptera made galls on leaves, flowers, stem or roots, and one Diptera and one Lepidoptera were leaf miners. The others were Schinia nundina (Drury), a flower and seed feeder, Craspedolepta veaziei (Patch), a sapsucker, Sparganothis distincta (Walsingham), a leaftier and Ophraella sexvittata (LeC.), a leaf chewer,. Biological information on the latter two insects can be found in Fontes (1985). The others are discussed below. The black or white leaf blister galls of Asteromyia carbonifera (Osten Sacken) were found frequently on the four goldenrod species during this study. The number of galls per leaf varied from one to many and sometimes covered the entire leaf. The relationship between A. carbonifera and the fungus that inhabits its gall was discussed by Batra (1964). The galls were caused by both fungus and insect activity. Asteromyia carbonifera associates with leaves of goldenrod already infected by the fungus Scle- rotium asteris (Schw.). Two or three midge larvae developed between layers of the fungus which forms a stroma on either side of the larval chamber. Weis (1982) showed that the formation of this stroma is an important mechanism for protection from the parasite Torymus capite (Huber). Also, the gall midge larvae were frequently parasitized by Tetrastichus sp. 1, T. homeri (Girault), and T. tesserus Burks, (Eulophidae). Another leaf blister gall found occasionally on leaves of goldenrod was caused by a midge identified as Asteromyia sp. nr. carbonifera (Osten Sacken). The circular galls are greenish, often surrounded by a purplish necrotic area. The midges were heavily parasitized by a eupelmid, probably Anastatus sp. Asphondylia monacha Os- ten Sacken made galls in developing buds. It established itself between the surface of the leaves while they were still in the bud and caused the adjacent tissues to form an oval cell between the two surfaces. The leaves continued their normal development but attached to each other where the gall developed. A common parasite of this species was Galeopsomyia haemon (Walker) (Eulophidae). Similar damage was caused by a midge (Dasyneura sp.) on the four goldenrod species, although the only injury observed was to the developing, opposed leaves of the growing bud. No adults were reared from the reddish larvae, so the species could not be determined. Another gall midge, Rhopalomyia solidaginis (Loew) attacked buds, transforming them into globular masses of deformed leaflets. In the center of the mass, a cylindrical chamber with tapered apex sheltered the yellowish larva. This gall was seen occasionally on all goldenrod species except S. canadensis var. scabra. It was also parasitized by G. haemon. Rhopalomyia new sp. was collected occasionally on S. fistulosa. The larvae developed inside individual globular galls with a tapered apex. These galls developed together forming an agglomeration on the stem. The parasitic wasp Torymus sp. nr. duplicatus (Hübner), probably a new species, was reared several times from this gall. The only flower gall observed in this study was made by the midge Schizomyia racemicola (Osten Sacken). It produced a rounded gall with tapered apex (frequently reddish) on the racemes of S. canadensis var. scabra and S. fistulosa. The larvae were orange-red and left the gall when disturbed. Gagne (1989) provides keys and descriptions for most of the cecidomyiids on Solidago in North America. The conspicuous galls of Gnorimoschema gallaesolidaginis (Riley) were occasionally seen on the stems of S. fistulosa and S. gigantea. A complete description of the biology of this insect is given by Leiby (1922). It is widely distributed and has also been reported from S. canadensis var. scarbra, S. nemoralis Aiton, and S. serotina Retz. Fully grown galls were found from July through the winter in Gainesville. In North Carolina, this insect hibernates in the egg stage (Leiby 1922), but in Florida it apparently overwin- 220 Florida Entomologist 77(2) June, 1994 ters as a pupa inside the gall. Two adults emerged from screened galls in February, just when goldenrod seeds were sprouting. The rarest gall collected was made on the roots by Eurosta prob. comma (Wied.). A large white maggot developed inside an elliptical, potato-like gall on the rhizomes of S. fistulosa. No adults of this species were obtained. The leaf mines of the gracillariid moth, Cremastobombycia solidaginis, were very common on the underside of leaves of S. canadensis var. scabra, S. fistulosa and S. gigantea. They were found from early May to early October. The tiny larva made an irregular, roundish blotched mine, usually centered on the underside of the leaves. As the larva grew the mine became elongate. By pupation, the leaf was folded in the damaged region and the mine became wrinkled. The elongate, white cocoon in which the larva pupated was suspended inside the fold by silken threads. This is a common species in the United States (Braun 1908). Bucculatrix solidaginiella Braun is a lyonetiid moth whose larvae feed in the growing tips or mine the leaves of various species of Solidago (Braun 1963). Only pupae and adults of this species were collected from the leaves of S. canadensis var scabra. The brightly colored yellowish larvae of the noctuid Schinia nundina were well concealed in the flower heads of S. fistulosa, where they fed on the developing seeds. Adults of this species occur from late July to late September in central and eastern U.S. (Forbes 1948). No adults were collected and the larvae were observed in October and early November. The jumping plant-louse, or psyllid, Craspedolepta veaziei has been reported from Solidago sp. (Caldwell 1938) and Aster (Crawford 1914). Adults were occasionally collected on leaves of S. fistulosa and S. gigantea from July to No- vember. Eight of the species recorded as feeding only on Solidago and Aster could be considered for introduction into Europe or Japan for the biological control of goldenrods: one attacks roots, Eurosta prob. comma; two are leaf chewers, Ophraella sexvittata and Sparganothis distincta; two are leaf miners, Agromyzidae sp. 1 and Cremasto- bombycia solidaginis; one is a leafgaller, Asteromyia carbonifera; and two attack flowers and seeds, Schizomyia racemicola and Schinia nundina. Before release, these insects would have to be tested to make certain that they do not feed on any cultivated Aster or other Asteraceae.

ACKNOWLEDGMENTS

We are grateful to the specialists at the University of Florida, the Division of Plant Industry of the Florida Department of Agriculture and Consumer Services, and the USDA Systematic Entomology Laboratory for their assistance in identifying the insects and plants. Voucher specimens have been placed in the Florida State Col- lection of Arthropods. Florida Agric. Expt. Sta. J. Series No. 7861.

REFERENCES CITED

ALTIERI, M. A. 1979. The design of pest stable corn agroecosystems based on the ma- nipulation of insect populations through weed management. Ph.D. Disserta- tion. Univ. of Florida, Gainesville. 68 pp. BATRA, L. R. 1964. Insect-fungus blister galls of Solidago and Aster. J. Kansas Ento- mol. Soc. 37: 227-234. BORROR, D. J., D. M. DELONG, AND C. A. TRIPLEHORN. 1976. An introduction to the study of insects, 4th ed. Holt, Rinehart and Winston, New York. 852 pp. BRAUN, A. F. 1908. Revision of the North American species of the genus Lithocolletis Hübner. Trans. Amer. Entomol. Soc. 34: 269. BRAUN, A. F. 1963. The genus Bucculatrix in America North of Mexico (Microlepi- doptera). Mem. Amer. Entomol. Soc. 18: 208. CALDWELL, J. S. 1938. The jumping plant-lice of Ohio. Ohio Biol. Survey Bull. 5: 241-242. Godfrey et al.: Establishment of B. afﬁnis on Hydrilla 221

CAPEK, M. 1971. The possibility of biological control of imported weeds of the genus Solidago L. in Europe. Acta Inst. For. Zvolensis: 429-441. CRAWFORD, D. L. 1914. A monograph of the jumping plant-lice or Psyllidae of the new world. Bull. U. S. Nat. Mus. 85: 182 pp. FELLER-DEMALSY, M. J., AND Y. Lamontagne. 1979. Pollen analysis of honeys from Quebec. Apidologie 10: 313-339. FONTES, E. M. G. 1985. The diversity of the insect fauna of four species of Solidago (goldenrods) in Gainesville and its relation to the plant architecture. Ph.D. dissertation. University of Florida, Gainesville. 120 pp. FORBES, W. T. M. 1948. Lepidoptera of New York and neighboring states. Part III. Cornell Univ. Agric. Exp. St., Ithaca. 188 pp. GAGNE, R. J. 1989. The plant-feeding gall midges of North America. Cornell Univ. Press. Ithaca, NY 356 p. HENSEL, M. 1982. Goldenrod. American Hort. 61: 14-25, 34. LEIBY, R. W. 1922. Biology of the goldenrod gall-maker Gnorimoschema gallaesolidaginis Riley. J. New York Entomol. Soc. 30: 81-94. LESAGE, L. 1986. A taxonomic monograph of the nearctic Galerucine genus Ophraella Wilcox (Coleoptera: Chrysomelidae). Mem. Entomol. Soc. Canada 133. METCALF, C. L., AND W. P. FLINT. 1951. Destructive and useful insects, 3rd ed. McGraw-Hill Book Co., Inc., New York. 1071 pp. NORBI, R. J., AND T. T. KOZLOWSKI. 1980. Allelopathic potential of ground cover species on Pinus resinosa seedlings. Plant and Soil 57: 363-374. TODD, J. W., AND D. C. HERZOG. 1980. Sampling phytophagous Pentatomidae on soybean, pp 438-478 in M. Kogan and D. C. Herzog [eds], Sampling methods in soybean entomology. Springer-Verlag, New York. 587 pp. WEIS, A. E. 1982. Use of symbiotic fungus by the gall maker Asteromyia carbonifera to inhibit attack by the parasitoid Torymus capite. Ecology 63: 1602-1605. WERNER, P. A., J. K. BRADBURY, AND R. S. GROSS. 1980. The biology of Canadian weeds. 45. Solidago canadensis L. Canadian. J. Plant Sci. 60: 1393-1409.

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OVERWINTERING AND ESTABLISHMENT POTENTIAL OF BAGOUS AFFINIS (COLEOPTERA: CURCULIONIDAE) ON HYDRILLA VERTICILLATA (HYDROCHARITACEAE) IN NORTHERN CALIFORNIA

K. E. GODFREY1, L. W. J. ANDERSON 1, S. D. PERRY2, AND N. DECHORETZ3 1USDA, ARS, Aquatic Weed Control Research Laboratory, University of California, Davis, CA 95616

2Aquatics Unlimited, 2150 Franklin Canyon Road, Martinez, CA 94553

3California Department of Food and Agriculture, 1220 N Street, Sacramento, CA 94271

ABSTRACT

Bagous affinis Hustache (Coleoptera: Curculionidae) has potential as a biological control agent for subterranean turions (also called tubers) of hydrilla (Hydrilla verticillata (L.f.) Royle; Hydrocharitaceae). The ability of B. affinis to overwinter and establish was investigated at 2 sites in northern California; a pond in Calaveras County and the Chowchilla River in Madera County. In cage studies conducted at the pond, B. affinis survived the winter for 2 seasons. After releases at the pond, B. affinis successfully reproduced and survived during the summer of 1992. Weevil larvae had damaged both sentinel and native tubers at the site. In the spring, following the

This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.uﬂ.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

222 Florida Entomologist 77(2) June, 1994 release of B. affinis at the pond, a light trap and baited shelter traps were used to detect weevils that had overwintered naturally at the site. No weevils were recovered. At the Chowchilla River site, B. affinis failed to establish, possibly due to a lack of tubers in the section of the river where the releases were made. Key Words: Aquatic weed control, biological control, field biology, noxious weeds.

RESUMEN

Bagous affinis Hustache (Coleoptera: Curculionidae) tiene potencial como agente de control biológico, al atacar los tallos subterraneos (tubérculos) de la Elodea de Florida (Hydrilla verticillata (L.f.) Royle; Hydrocharitaceae). La habilidad de B. affinis para invernar y establecerse fué investigada en dos sitios del norte de California: un estanque en el condado de Calaveras y el río Chowchilla en el condado de Madera. En estudios en jaulas, llevados a cabo en el estanque, B. affinis sobrevivió el invierno a lo largo de dos estaciones más. Después de las liberaciones en el estanque, B. affinis se reprodujo exitosamente y sobrevivió durante el verano de 1992. Las larvas del picudo dañaron tanto los tuberculos secundarios como los tuberculos primarios. En la primavera, luego de la liberación de B. affinis, se utilizó una trampa de luz y trampas con cebo para detectar los picudos que habían imvernado en el estanque. No se recuperaron picudos. En el río Chowchilla B. affinis no se estableció, posiblemente debido a la falta de tubérculos en la sección del río donde se efectuaron las liberaciones.

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Hydrilla verticillata (L.f.) Royle (Hydrocharitaceae) has been a serious weed in ponds, lakes and waterways since its discovery in California in 1976 (California De- partment of Food and Agriculture 1991). As a part of a state-mandated eradication program designed to reduce the spread and density of H. verticillata (hydrilla), infested aquatic sites are quarantined, or closed to fishing and recreation. The hydrilla then is treated with herbicides, physically removed by dredging, and/or controlled by release of sterile triploid grass carp, Ctenopharyngodon idella (Cuvuer and Valenci- ennes; Cyprinidae). The fish have only been released in canals in the Imperial Irri- gation District. These tactics are very effective against the above-ground portions of the plant, however, subterranean vegetative propagules, called subterranean turions or tubers, have a greater survival rate when using these tactics than the above- ground portions of the plant. Tubers are the primary cause of reinfestations. Cur- rently, densities of hydrilla tubers are reduced by attrition, use of soil fumigants, or dredging. Chemical and physical methods are often disruptive to the aquatic environment. Thus, in an attempt to reduce disruption to wetland environments, biological control of hydrilla tubers was investigated. In the 1970's and 1980's, a search was initiated for organisms that fed upon hydrilla tubers on the Indian subcontinent (Baloch et al. 1980). A weevil, Bagous affinis Hustache (Coleoptera: Curculionidae), was found feeding on hydrilla tubers in central Pakistan (Baloch et al. 1980). The weevil had infested 90-100% of the exposed hydrilla tubers as waters receded during the dry season. No regrowth of hydrilla occurred at this site in the following wet season (Baloch et al. 1980). Bagous affinis was imported into quarantine in the United States in the mid 1980's to study its biology and host plant range (Buckingham 1988). The life cycle in the laboratory was determined, and it was found that the weevil could not withstand submergence [not longer than 2.5 days (larvae), and not longer than 5 days (adults); Bennett and Buck- ingham 1991]. From studies of host plant range, B. affinis was found to feed and complete development only on hydrilla (Buckingham 1988). B. affinis was imported into California in the summer of 1991 to determine if it could be used to reduce densities of hydrilla tubers biologically. The weevil seemed

223 well suited to conditions in northern California because in some of the water systems infested with hydrilla, drawdown conditions exist naturally or may be created artiﬁ- cially. The suitability of B. afﬁnis as a biological control agent for hydrilla tubers in California was investigated by examining its ability to overwinter and to establish after release.

MATERIALS AND METHODS

All B. affinis used were the progeny of weevils collected outside Bangalore, India, in April 1991. The weevils were cultured in quarantine at the Florida Biological Con- trol Laboratory, Gainesville, Florida, for 1 generation before shipment of the weevils to California. The weevils were maintained in laboratory culture at the USDA Aquatic Weed Control Research Laboratory, Davis, California, for 2-10 generations before use in the experiments. Voucher specimens were deposited at the Bohart Mu- seum, Department of Entomology, University of California, Davis, California. The ability of B. affinis to overwinter in northern California was determined for 2 consecutive fall-spring seasons with caged weevils placed along the bank of a hydrilla-infested pond near Mountain Ranch, California, in Calaveras County (approxi- mate lat. 38oN, long. 120oW). In the first fall-spring season, the studies were initiated by placing 35 cages (28.5 cm diam x 35.5 cm height) at the pond on 23 Sep- tember 1991. Each cage contained 10-15 cm of soil in which 50 hydrilla tubers had been buried at a depth of about 5 cm. Only dioecious hydrilla tubers were used in these experiments because dioecious hydrilla is the predominant type of hydrilla found in California. On the soil surface, approximately 20 hydrilla stems covered a 2- cm cube of water-saturated floral foam. The hydrilla stems served as a food source, and the floral foam, an oviposition substrate. [Floral foam was found to be an accept- able substrate for oviposition, and more convenient to work with in field studies than water-soaked wood, the natural oviposition substrate (Bennett & Buckingham 1991).] Ten B. affinis adults were placed in each cage. Five randomly selected cages were returned to the laboratory on the following dates: 15 October, 1 November, and 18 December 1991; 7 February, 3 March, 9 April, and 6 May 1992. On each sample date, fresh hydrilla stems were added to the cages remaining in the field. The contents of the retrieved cages were sorted by examining the hydrilla stems to detect B. affinis adults, and the soil was washed through a 5-mm sieve to recover tubers. The tubers were examined for evidence of feeding by B. affinis larvae and to recover any B. affinis. The number and life stage of any B. affinis, and the number and condition (i.e., fed upon or not fed upon) of the tubers were recorded. In the second fall-spring season, the 35 cages were placed at the pond on 21 Sep- tember 1992. The contents of each cage was similar to that in the first season, except that 25 tubers, and 5 B. affinis adults were placed in each cage. In addition, each cage contained two 20-ml vials filled with water and stoppered with sponges. Food and water were replenished every 2 weeks for the first 2 months. For the remainder of the study, fresh food and water were provided on the following sampling dates: 19 October, 16 November, and 14 December 1992; 19 January, 17 February, 15 March, and 19 April 1993. The procedures for sorting the contents of the cages were the same as those used in the first season. The trend in percent of tubers fed upon by B. affinis was regressed against time for both years using PROC REG (SAS Institute 1982). During both seasons, the air and soil temperatures within the cages were moni-  tored using thermistors connected to a Li-Cor LI-1000 Data Logger (Lincoln, NE). Within the cage, the thermistor for the air temperature was positioned approximately 10 cm above the soil surface, and that for the soil, buried approximately 5 cm below the soil surface. The hourly mean air and soil temperatures were recorded.

224 Florida Entomologist 77(2) June, 1994

The establishment potential of B. affinis was investigated at 2 sites in northern California. The first site was the pond in Calaveras County used for the overwintering studies. Releases of B. affinis eggs and adults were made in 1991 and 1992, respectively. The other site was along the Chowchilla River near Raymond, California, in Madera County (approximately lat. 37oN, long. 120oW) where adult weevils were released in 1991 only. For releases at the pond, the water level had to be drawn down, and this was done from 21-26 August 1991 and from 18-21 May 1992. Draw- down was necessary to insure access to tubers by B. affinis larvae. On 30 August 1991, the pre-release tuber density in the exposed pond bottom was estimated by taking three 15-cm diam core samples (sample depth = 10 cm) from each of 15 transects that were arranged in a spoke-like fashion around the pond (total number of samples = 45). Each transect ran from the edge of the water to the bank of the pond. The first core in each transect was taken next to the edge of the water, the second, 1.5 m from the edge of the water, and the third, 3 m from the edge of the water. On 8 September 1992, after all B. affinis releases had been completed, the post-release density of tubers was estimated as described above except that 25 transects were sampled (total number of samples = 75). The area of exposed pond bottom sampled for the post-release estimate was approximately the same as that for the pre-release estimate of tuber density. Soil cores were bagged, returned to the laboratory, and washed through a 5-mm sieve. The number and condition (i.e., fed upon or not fed upon) of the tubers were recorded. The mean pre- and post-release densities were compared using a t-test (Steel &Torrie 1960). The first set of releases at the pond was made on 24 September 1991 and consisted of 3 releases of 50 B. affinis eggs each. The eggs were placed near the edge of the water at sites thought to have the greatest probability of having tubers. At each site, pieces of water-soaked wood containing eggs were placed on the surface of the soil, and covered with a 5-cm layer of moist sphagnum moss. The release site was covered with a wire-mesh cage (mesh size 2.5 x 3.5 cm; cage diam 75 cm, height 1.2 m) to exclude cattle that used the pond for water. On 15 October 1991, the soil around (diam of area removed = 40 cm) and under (depth = 10 cm) the release site was placed in a plastic bag and returned to the laboratory. The soil was washed through a 5-mm sieve to recover tubers, and the number and condition of any tubers recovered were recorded. In the second set of releases, 500 B. affinis adults were released on the following dates: 21 May (200), 17 June (100), 15 July (100), and 11 August 1992 (100). On each release date, the weevils were divided among 4 randomly chosen sites near the edge of the water. At each release site, weevils were provided hydrilla stems, a 2-cm cube of water-saturated floral foam, and a source of free water. To monitor the success of each release, we buried a mesh bag (1 x 1 cm mesh) containing 25 tubers 5 cm below each release site. These tubers were used as “sentinel” tubers to monitor feeding ac-  tivity by B. affinis larvae. The release site was covered with a screen made of Saran screening (Lumite-Synthetic Industries, Gainesville, GA; 32 x 32 mesh) to keep birds and other small animals out of the release site. The sentinel tubers were recovered 25-30 days after release. The tubers were broken in half to determine the presence of B. affinis. The number and life stage of any B. affinis found, and the number and condition of the sentinel tubers were recorded. During the releases, the mean hourly ambient air temperature and soil temperature (5 cm below the soil surface) were  monitored using thermistors connected to a Li-Cor LI-1000 Data Logger (Lincoln, NE). To determine if any B. affinis produced as a result of the releases in 1992 survived the winter, 2 types of traps, black light and baited shelter trap, were deployed around the pond during the spring of 1993. The black light (UV, 365 nm) trap, suspended by a frame 1.5 m above the soil surface on the bank of the pond, was used to attract emerging weevils in “flight mode” (i.e., energy resources directed toward the

225 development of indirect flight muscles; Bennett & Buckingham 1991). The insects attracted to the light were collected in a jar (1 liter) containing a 1:1 mixture of ethyl- ene glycol and water. The trap was operated for 3 consecutive days per week beginning 19 April and ending 26 May 1993. The contents of the jar were sorted under 10X magnification. The baited shelter trap was used to attract weevils that were in “reproductive mode” (i.e., energy resources directed toward the reproductive system, not the indirect flight muscles; Bennett & Buckingham 1991). Each shelter trap consisted of a 1 liter container (diam = 12 cm; height = 12 cm) that contained soil and 25 tubers buried 5 cm below the soil surface. A 2-cm cube of water-saturated floral foam and a mesh bag (1 x 1 cm mesh) containing hydrilla stems were placed on the soil surface. Each trap was covered with a screen (2.5 x 3.5 cm mesh) to exclude other animals. Each week of light trapping, 5 baited shelter traps were buried in the soil around the light trap such that the top rim of the container was flush with the surrounding soil surface, much like a pitfall trap. The shelter traps were left in the field for 1 week. Upon return to the laboratory, the hydrilla stems from the tops of the traps were examined to recover any adult weevils. The soil within the trap was covered with moist sphagnum moss, and the traps were held for 30 days under standard rearing conditions [27oC, 14:10 (L:D)] to allow any B. affinis immatures to develop to the adult stage. The contents of the traps were examined, and the number of B. affinis recovered was recorded. The second B. affinis release site in California was along the Chowchilla River. This site was selected because the river goes through a natural low water period. Most of the river bed was exposed from mid summer through early fall. The density of hydrilla in the river was low because the river had been treated with herbicides for the past 5 years in an attempt to eradicate hydrilla. The release site was a small pool that had a few hydrilla plants and tubers. The density of hydrilla tubers within the pool could not be quantitatively estimated without destroying the site. Bagous affinis adults were released on 11 October (200) and 25 October 1991 (103). The site was left undisturbed until 12 November 1991 when the site was sampled for tubers. Stan- dard tuber sampling (i.e., using a 15-cm diam core sampler) was attempted, but abandoned because the soil in the bottom of the pool was a very fine sand that did not hold together as the sampler was pulled up through the 60 cm of water covering the site. Instead, a shovel was used to remove soil from the bottom of the pool. The soil was washed through a 5-mm sieve to recover tubers. The number and condition of tubers recovered were recorded. To determine if B. affinis had overwintered and established at the Chowchilla River site, we used black light trapping and tuber sampling. Black light trapping (as described above) was done continuously from 23 - 27 April and from 30 April - 4 May 1992. The ambient air temperature at the site was recorded during trapping with a recording thermometer (PTC Instruments Los Angeles, CA). The river bed was sampled during the summer, the natural low water period. If B. affinis had established at this site, then feeding damage should have been evident on the tubers. Tubers were sampled by taking fifty 15-cm diam core samples (depth of sample = 10 cm) on 9 July and 21 August 1992 (total number of samples = 100) from a 30-m section of the river. The midpoint of this section of river was the release site. Transects traversed the river bed at 3 m intervals beginning at the release site and running up and down river. Five cores were taken from the release site and from each transect. The core samples were washed through a 5-mm sieve to recover tubers. The number and condition of the tubers recovered were recorded.

226 Florida Entomologist 77(2) June, 1994

RESULTS

Bagous affinis survived the winter in cages at the pond in Calaveras County in 1991 - 1992 and in 1992 - 1993 (Table 1). In the first season, 2 adults were recovered on the 9 April 1992 sampling date (Table 1). These individuals were in all likelihood the F1 progeny of the adults originally placed within the cages because the mean number of tubers fed upon and the percent of tubers fed upon increased with the length of time the cages were in the field (Table 1). Linear regression of the percent of tubers fed upon against time resulted in a significant positive regression (F = 54.2; df = 1,5; P < 0.05; R2 = 0.92). Much of this feeding damage was characteristic of that produced by B. affinis larvae. This trend in feeding activity suggested that B. affinis was successfully reproducing within the cages. In addition, in the laboratory under optimal conditions, B. affinis adults survive a mean of 127.5 days (range 55 - 225 days; Bennett &Buckingham 1991). The 9 April 1992 sampling date was 231 days after the cages were placed in the field. In the second season, 4 adults were recovered, 3 from the samples taken on 17 February, and 1 from the 15 March 1993 sampling date (Table 1). These individuals also were in all likelihood the progeny of adults originally placed within the cages because as in the first season, the percent of tubers that had been fed upon increased with the length of time the cages were in the field (Table 1). Linear regression of the percent of tubers fed upon against time resulted in a significant positive regression (F = 40.3; df = 1,5; P < 0.05; R2 = 0.89). In addition, the 17 February and the 15 March sampling dates are 150 and 177 days after the initial placement of the cages in the field, respectively. The 3 adults recovered from the 17 February sampling date began ovipositing about 7 days after being brought into the laboratory and produced 44 progeny when held under rearing conditions. These results show that B. affinis can successfully reproduce after overwintering in northern California. The mean number of tubers recovered in both seasons decreased with the length of time the cages were in the field (Table 1). This decrease was due to the degradation of tubers either naturally or after having been fed upon by B. affinis larvae. Van & Steward (1990) found that some tubers will degrade naturally through time. The reason for this degradation is not known. In the degradation of tubers through the feeding activity of B. affinis, the larvae provide an opening for other organisms to en- ter and attack the tuber. In some cases, entire tubers were destroyed, whereas in others, whole tubers and pieces of tubers were recovered.

TABLE 1. TOTAL NUMBER OF B. AFFINIS RECOVERED, THE MEAN NUMBER (STD. DEV.) OF TUBERS RECOVERED, THE MEAN NUMBER (STD. DEV.) OF TUBERS FED UPON BY B. AFFINIS LARVAE, AND THE PERCENT OF TUBERS FED UPON FROM THE OVERWINTERING STUDIES CONDUCTED AT THE POND IN CALA- VERAS COUNTY IN 1991-1992 AND 1992-1993. THE MAXIMUM NUMBER OF TUBERS RECOVERABLE WAS 50 IN 1991-1992 AND 25 I N 1992-1993.

Mean Mean No. of No. of No. of Percent B. afﬁnis Tubers Tubers of Tubers Date Recovered1 Recovered Fed Upon Fed Upon 1991 - 1992 15 October 2A 48.2 3.2 6.6 (1.64) (2.17) 1 November 1A 37.8 4.2 11.1 (7.69) (2.68) 18 December 1A 32.0 8.6 26.9 (15.87) (2.19) 7 February 0 26.8 7.4 27.6 227

TABLE 1.(CONTINUED) TOTAL NUMBER OF B. AFFINIS RECOVERED, THE MEAN NUM- BER (STD. DEV.) OF TUBERS RECOVERED, THE MEAN NUMBER (STD. DEV.) OF TUBERS FED UPON BY B. AFFINIS LARVAE, AND THE PERCENT OF TUBERS FED UPON FROM THE OVERWINTERING STUDIES CONDUCTED AT THE POND IN CALAVERAS COUNTY IN 1991-1992 AND 1992-1993. THE MAXIMUM NUMBER OF TUBERS RECOVERABLE WAS 50 IN 1991-1992 AND 25 I N 1992- 1993. Mean Mean No. of No. of No. of Percent B. afﬁnis Tubers Tubers of Tubers Date Recovered1 Recovered Fed Upon Fed Upon (10.33) (3.29) 3 March 0 23.8 7.2 30.3 (13.61) (4.92) 9 April 2A 17.8 9.4 52.8 (5.79) (7.92) 6 May 0 28.4 14.4 50.7 (12.36) (6.31) 1992 - 1993 21 October 2L 18.8 10.4 55.3 (4.21) (3.21) 16 November 1L 16.4 9.8 59.8 (4.51) (2.86) 14 December 1A 9.4 6.6 70.2 (3.05) (2.97) 19 January 0 10.4 8.2 78.9 (5.37) (4.15) 17 February 3A 4.6 4.4 95.7 (2.61) (2.51) 15 March 1A 3.8 3.6 94.7 (2.39) (2.30) 19 April 0 2.6 2.4 92.3 (4.77) (4.34)

1L = larva, A = Adult.

The soil and air temperatures within the cages were similar for both fall-spring seasons. At no time during these time periods did the soil temperatures within the cage drop below 1oC or rise above 29oC. The air temperatures within the cages were more variable. The lowest air temperatures ranged from -8.5 to -13.2oC in mid-winter, and the highest air temperatures ranged from 43.6 to 45.3oC in the fall. Releases of B. affinis at the pond resulted in successful reproduction and survival by B. affinis in the summer of 1992. For the first set of releases in which B. affinis eggs were released, establishment of the weevil could not be monitored because no tubers were found at the release sites. For the second set of releases, however, sentinel tubers were used to monitor establishment. From the weevils released on 21 May 1992, 5 pupae and 2 adults were recovered from sentinel tubers (Table 2). For the other release dates, no weevils were recovered from the sentinel tubers, however, some tubers had been fed upon by B.affinis larvae (Table 2). The air and soil (5 cm depth) temperatures during the releases were in a range suitable for B. affinis reproduction and development. 228 Florida Entomologist 77(2) June, 1994

To further investigate the establishment of B. affinis at this site, the density of native tubers was estimated before and after releases of B. affinis. Before the release of any B. affinis, the native tuber density within the pond was estimated as 0.82 tubers per 15-cm core (Table 3). After all releases were complete, the native tuber den-

TABLE 2. THE MEAN NUMBER (STD. DEV.) OF SENTINEL TUBERS RECOVERED, THE MEAN NUMBER (STD. DEV.) OF SENTINEL TUBERS FED UPON, AND THE TOTAL NUMBER OF B. AFFINIS RECOVERED FROM SENTINEL TUBERS FROM RE- LEASES OF B. AFFINIS ADULTS MADE AT A POND IN CALAVERAS COUNTY IN 1992. Mean No. of Mean No. of Sentinel Sentinel Total Sampling Tubers Tubers B. afﬁnis Date Recovered Fed Upon Recovered1 21 May 23.0 6.0 5P,2A (0.82) (2.71) 17 June 24.332 1.33 0 (1.15) (0.58) 15 July 24.5 2.75 0 (0.58) (1.26) 11 August 25.0 0.75 0 (0.82) (0.96)

1P = pupa, A = adult. 2One release site destroyed by cattle. Maximum total recoverable sentinel tubers = 75.

sity within the pond was estimated as 0.12 tubers per 15-cm core (Table 3). This apparent decrease in native tuber density was not statistically significant (P > 0.05). However, 22% of the native tubers recovered had feeding damage that resembled that imparted by B. affinis larvae. We assume that this feeding damage had been caused by B. affinis because no native organisms are known to feed inside hydrilla tubers in the United States (Balciunas & Minno 1985). Light and baited shelter traps operated in the spring of 1993 did not capture any overwintering progeny from B. affinis released at the pond in the summer of 1992. However, 6 of the 125 tubers (4.8%) examined in the last set of baited shelter traps (placed in the field 24 May 1993), had damage resembling that caused by B. affinis larvae. Releases of B. affinis made along the Chowchilla River did not result in establishment of the weevil. Six tubers were recovered from samples collected approximately 30 days after release. Of these tubers, 2 had damage characteristic of B.affinis larval feeding, suggesting that the weevils initially released at this site were successful in reproducing. In subsequent monitoring of the site, no weevils were captured by the light trap, and no tubers (i.e., tuber density = 0 tubers per 15 cm-core) were recovered in the tuber sampling of the river. The absence of hydrilla tubers in this section of the river was due to hydrilla plant removal and treatment of the river with herbicides in previous years. Given the lack of tubers in this section of the river, it seems unlikely that B. affinis could have survived without food resources even if it could overwinter at the site. 229

TABLE 3. THE MEAN NUMBER (STD. DEV.) OF HYDRILLA TUBERS RECOVERED IN 15-CM CORE SAMPLES FROM THE EXPOSED POND BOTTOM IN CALAVERAS COUNTY. Transects Overall Water’s 1.5 m from 3 m from Mean No. Sampling Date Edge Water’s Edge Water’s Edge of Tubers 30 August 1991 0.13 0.40 1.93 0.82 (0.35) (0.83) (2.60) (10.67) 8 September 1992 0.12 0.16 0.08 0.12 (0.44) (0.47) (0.28) (3.52)

DISCUSSION

Bagous affinis demonstrated an ability to adapt to the climate in northern Cali- fornia. Adult weevils were able to survive the winter within cages despite soil temperatures dropping to near 1oC and air temperatures approaching -14oC. These cold temperatures did not appear to reduce the reproductive success of the weevils because weevils recovered from overwintering cages produced progeny in numbers equivalent to those weevils in the colony when placed under normal rearing conditions. The ability of the weevils to survive the winter within the cages suggested that the weevils may also be able to overwinter naturally. However, no naturally-overwintering weevils were trapped at either release site in light traps. This is not surprising considering that the greatest attraction to a light trap by a particular species of insect occurs within a narrow range of environmental conditions. For B. affinis, the environmental conditions for maximum attraction to a light trap include temperatures at sunset at or above 20oC and little or no wind (Buckingham et al. 1994). For both release sites, the temperature at sunset was at or above 20oC for about half of the trapping days; however, on all trapping days light to moderate winds prevailed. In addition, for B. affinis to be attracted to the light trap, the weevils would have to be in “flight mode”, not “reproductive mode”. The density of weevils that may have overwintered naturally also would not have been large, thereby making it difficult to detect weevils using light traps. The baited shelter traps should have been more attractive to any naturally-overwintering B. affinis that were in reproductive mode than the light traps. No weevils were recovered from the traps, but a small amount of feeding damage was detected in the tubers from the traps. This feeding damage suggests that the density of overwintering weevils was probably very low. Given its demonstrated ability to overwinter, it is not surprising that the weevil successfully reproduced using native tubers at 1 of the 2 release sites. Its failure to establish at the Chowchilla River site was probably due to the lack of tubers in the part of the river where it was released. In order for B. affinis to establish, it must have a food resource. At the pond in Calaveras County, the weevil reproduced successfully and survived through the summer of 1992. Evidence of its reproduction and survival was found in both sentinel and native tubers. In the sentinel tubers, B. affinis immatures were recovered after the first release, and tubers damaged by B. affinis were recovered in subsequent releases (Table 2). The lack of recovery of B. affinis in the later releases can be explained by the developmental rate of B. affinis. For the first set of releases, the soil temperature at 5 cm (the depth of the sentinel tubers) was near 21oC. At 21oC, the development of B. affinis from larva to adult requires approximately 30 days (K.E.G., unpublished data). Given a short amount of time for oviposition and egg hatch, it was expected that the sentinel tubers recovered about 30 days after release would contain B. affinis immatures. For the remaining re- 230 Florida Entomologist 77(2) June, 1994 leases, the soil temperature at 5 cm increased to 25 - 28oC. The development of B. affinis from larva to adult requires approximately 20 and 12 days at 25 and 28oC, respectively (K.E.G., unpublished data). Again, given a short period of time for oviposition and egg hatch, it was expected that B. affinis could have completed development leaving only damaged tubers. Of the native tubers recovered from the pond after all B. affinis releases were complete, 22% had been fed upon by B. affinis. This level of tuber damage was within the range of tuber damage found by Buckingham et al. (1994) at a lake in Florida where B. affinis had successfully survived and reproduced for 3 months. They found from 0.57 - 32% of the tubers recovered from the lake had been fed upon by B. affinis. In addition, they found degradation of the tubers after B. affinis feeding as we did in this study. Bagous affinis has potential as a biological control agent for hydrilla tubers in California. This weevil could be used as one element in the long-term management strategy to reduce numbers of hydrilla tubers. To do this, B. affinis should be released at sites that undergo annual drawdowns and have tuber densities sufficient to sustain populations of the weevil from year to year. In addition, B. affinis could also be used in more intensive, short-term management of hydrilla tubers through inun- dative releases at sites where more rapid reductions of tuber numbers are required.

ACKNOWLEDGMENT

We acknowledge S. Krishnaswamy for collecting and shipping the weevils from India; and T. Sankaran for arranging collection, packaging, and delivery of the weevils from India to Florida. We also acknowledge C. Bennett and G. Buckingham for quarantine handling and initial colonization of the weevils. We also acknowledge the assistance given by R. O'Connell, R. Ng, and F. Zarate in locating sites, in the maintenance of water levels at the pond and in tuber sampling. We thank D. Spencer, G. Buckingham, and C. Turner for reviewing an earlier draft of the manuscript. Re- search supported by a grant from the California Department of Food and Agricul- ture. Mention of a proprietary product does not constitute an endorsement or a recommendation for its use by USDA.

REFERENCES CITED

BALCIUNAS, J. K., AND M. C. MINNO. 1985. Insects damaging Hydrilla in the USA. J. Aquat. Plant Manage. 23: 77-83. BALOCH, G. M., SANA-ULLAH, AND M. A. GHANI. 1980. Some promising insects for the biological control of Hydrilla verticillata in Pakistan. Trop. Pest Manage. 26: 194-200. BENNETT, C. A., AND G. R. BUCKINGHAM. 1991. Laboratory biologies of Bagous affinis and B. laevigatus (Coleoptera: Curculionidae) attacking tubers of Hydrilla verticillata (Hydrocharitaceae). Ann. Entomol. Soc. America 84: 420-428. BUCKINGHAM, G. R. 1988. Reunion in Florida - Hydrilla, a weevil, and a fly. Aquatics 10: 19-25. BUCKINGHAM, G. R., C. A. BENNETT, AND E. A. OKRAH. 1994. Temporary establishment of the hydrilla tuber weevil (Bagous affinis) during a drawdown in north-central Florida. J. Aquat. Plant Manage. 32: (in press). CALIFORNIA DEPARTMENT OF FOOD AND AGRICULTURE. 1991. Hydrilla verticillata. Detection Manual. Rev. 10/11/91. 3pp. SAS INSTITUTE, INC. 1982. SAS User's Guide: statistics. 1982 Edition, Cary, NC. STEEL, R. G. D., AND J. H. TORRIE. 1960. Principles and procedures of statistics. McGraw-Hill Book Company, Inc. NY. VAN, T. K., AND K. K. STEWARD. 1990. Longevity of monoecious hydrilla propagules. J. Aquat. Plant Manage. 28: 74-76.

Sutton & Steck: Discrimination of Fruit Fly Larvae 231

DISCRIMINATION OF CARIBBEAN AND MEDITERRANEAN FRUIT FLY LARVAE (DIPTERA:TEPHRITIDAE) BY CUTICULAR HYDROCARBON ANALYSIS

BRUCE D. SUTTON AND GARY J. STECK Division of Plant Industry Florida Department of Agriculture and Consumer Services Gainesville FL 32614-7100

ABSTRACT

Larvae of the Caribbean fruit fly, Anastrepha suspensa (Loew) can be differenti- ated from those of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) with nearly 100% accuracy by analysis of cuticular hydrocarbon (CHC) patterns. A discriminant model is presented based on samples of feral and laboratory Caribbean and Mediterranean fruit flies. The difference in the ratio of two components is sufficient to discriminate third instars of the species. Accuracy in discrimination, utilization of specimens that are damaged, dried or otherwise unusable for morphometric, isozyme or DNA analysis, low cost per sample and automation of the process, all make CHC analysis a particularly effective solution for identification of these two species. Key Words: Anastrepha suspensa, Ceratitis capitata, gas chromatography, discriminant analysis.

RESUMEN

Las larvas de la mosca Caribeña de las frutas, Anastrepha suspensa (Loew) y las de la mosca del mediterraneo, Ceratitis capitata (Wiedemann) pueden ser diferencia- das entre sí con aproximadamente un 100% de precisión mediante análisis de los pa- trones de los hidrocarburos cuticulares. Se presenta un modelo para ambas moscas basado en muestras ferales y de laboratorio. La diferencia en la relación de dos com- ponentes es suﬁciente para discriminar terceros estadíos de cada especie. La pre- cisión en la discriminación, la posibilidad de trabajar con especímenes que se encuentran dañados, secos, o de alguna manera inutilizables para el análisis morfo- métrico, de isozimas o DNA, el bajo costo por muestra y la automatización del pro- ceso, hacen el análisis de hidrocarburos una solución particularmente efectiva para la identiﬁcación de estas dos especies.

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The state of Florida, as a major producer and exporter of fruits and vegetables, maintains a number of programs to deal with tephritid fruit fly pests. Anastrepha suspensa (Loew), the Caribbean fruit fly or caribfly, successfully invaded and rapidly colonized the southern half of the state during the 1960's (Clark & Weems 1989) ne- cessitating costly treatment and population monitoring programs to protect commercial crops from damage, and to certify fruit free of this insect for exportation (Riherd 1993). Additionally, state and federal agencies maintain a vigilant quarantine and trapping program to minimize introductions and provide early detection of other exotic fruit flies [e.g. Ceratitis capitata (Wiedemann), Mediterranean fruit fly or medfly, or other Anastrepha species]. Once new fruit fly introductions are detected, usually by capture of adult fruit flies in specially baited traps, a well-defined action plan must go into effect. Captured flies are identified and evaluated for reproductive condition, trap density in the vicinity of the original trap site is intensified, and fruits, if present, are checked for the presence of larvae. Detection of an active infestation of This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.ufl.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

232 Florida Entomologist 77(2) June, 1994 an exotic species and its focal point, i.e., fertile adults ovipositing into susceptible fruits and producing viable larvae, is critical before undertaking an effective action plan. Due to the widespread occurrence of A. suspensa, however, Florida faces a special problem in finding such focal points. In practical terms, we face a needle-in-a- haystack problem if the new detection occurs in an area where A. suspensa is already actively breeding. Accurate identification of fruit fly larvae is difficult to impossible given current knowledge (Steck et al. 1990). The problem in Florida is simplified to the extent that the identification is an either-or decision, i.e., A. suspensa or newly detected exotic. The third instar of A. suspensa has been described in useful detail by Heppner (1984) and is included in the key of Steck et al. (1990), but other immature stages of this species have not been adequately described. Immature stages of medfly also have been described (Phillips 1946, Hardy 1949). Usually it is possible to differ- entiate larvae of the two species using morphological criteria; however, the natural range of morphological variability includes overlap in character state distributions, and some specimens (especially earlier instars) cannot be reliably identified. About 10% of A. suspensa and C. capitata larvae fall into the indeterminate category [H.A. Denmark, Florida Department of Agriculture and Consumer Services (FDACS), pers. comm.]. Given the economic costs that may accrue from the mis-identification of fruit fly larvae, it is desirable that effective adjuncts to morphological identification of larvae be found and implemented. In this paper we describe the use of cuticular hydrocarbon (CHC) analysis to dif- ferentiate C. capitata and A. suspensa immature stages. Carlson & Yocom (1986) provided the groundwork for this analysis by describing CHC chromatographic patterns of six economically important fruit fly taxa including C. capitata and A. suspensa. The third instar larvae of A. suspensa and C. capitata were found to exhibit quantitatively different hydrocarbon patterns. More recently, Sutton & Carlson (1993) investigated CHC variation in larvae of A. fraterculus (Wiedemann), A. obliqua (Macquart), A. suspensa and other Anastrepha taxa and constructed discriminant models. Unfortunately, it is not possible to reconstruct realistic classification error estimates from the sample statistics as published, given the limited number of samples and the potential bias to which it could lead. Thus, it is necessary to sample the range of variation before an effective discriminant model can be constructed that not only distinguishes between A. suspensa and C. capitata larvae based upon CHC patterns, but also specifies robust classification error rates. Consequently, we have extended the previous analyses to include representative samples of third instar A. suspensa larvae from a number of host fruits in Florida and the Caribbean, and third instar C. capitata larvae from various hosts in Central America and Hawaii. A small number of second instar larvae was also analyzed.

MATERIALS AND METHODS

A total of 159 third instar larvae of A. suspensa was analyzed. The samples originated from eight feral collections in Dade County, Florida, taken from loquat (Erio- botrya japonica (Thumb.) Lindl.), calamondin (Citrofortunella x mitis Blanco), guava (Psidium guajava L.), cattley guava (Psidium cattleianum Sabine), Surinam cherry (Eugenia uniﬂora L.) and tropical almond (Terminalia catappa L.); one feral collection from Lake Co., Florida, from guava; and four collections from Puerto Rico from guava and tropical almond. Additional third instar larvae of A. suspensa were obtained from the USDA-ARS laboratory colony (Gainesville, FL) and the FDACS mass-rearing facility (Gainesville, FL), together with 12 second instar larvae. C. capitata samples comprised 99 specimens of third instar larvae taken from six feral collections in Hawaii originating from coffee (Coffea arabica L.), persimmon (Diospyros kaki L. f.), and Jerusalem cherry (Solanum pseudocapsicum L.); samples from the USDA-APHIS mass-rearing facility at Waimanalo and the USDA-ARS col-

Sutton & Steck: Discrimination of Fruit Fly Larvae 233 ony at Waimea; one feral collection from calamondin from Guatemala, and samples from laboratory colonies at USDA-APHIS and the Petapa mass-rearing facility. A total of 12 second instar larvae was obtained from the Waimanola facility and two feral second instars were collected from coffee in Hawaii. All specimens are vouchered with the Florida State Collection of Arthropods, FDACS, Gainesville, Florida. Individual whole larvae (fresh, freshly frozen, or dried) were extracted and prepared as previously described (Carlson & Yocom 1986, Sutton & Carlson 1993). GC analyses were made with a Tracor Model 540 Gas Chromatograph (Austin, TX) fitted with a nonpolar fused silica capillary column (BP-1, 50 m x 0.15 mm id, 0.25 µm film thickness, Scientific Glass Engineering, Austin, TX), cool on-column injector (OCI-3, Scientific Glass Engineering, Austin, TX) and Tracor flame-ionization detector. Puri- fied hydrogen was used as the carrier gas at a pressure of 80 psi providing a linear flow-rate of 35-45 cm/s; the oven was temperature programmed from 60-225oC at 20o per minute, and 225-320oC at 1.5o per minute. Areas were calculated for each of the selected peaks for each sample using the Nelson Model 2100 Chromatography Data System, Revision 5.0 (Nelson Analytical, Inc., Cupertina, CA). The SAS System, Version 6.01, (SAS Institute, Inc., Cary, NC) was utilized for the discriminant analysis and the density plot was constructed using Sygraph, Version 1.1 (SYSTAT, Inc., Evanston, IL).

RESULTS AND DISCUSSION

The CHC composition of larvae of A. suspensa and C. capitata is dominated by homologous series of n-alkanes and 2-monomethylalkanes with lesser quantities of other mono- and dimethylalkanes (Carlson & Yocom 1986, Sutton & Carlson 1993). The unsaturated hydrocarbons of A. suspensa larvae include squalene together with small quantities of as yet unidentiﬁed components (Sutton & Carlson 1993). Chro- matograms of third instar A. suspensa and C. capitata extracts (Figure 1) exhibit consistent quantitative differences in the relative proportions of 2-methyloctacosane at Kovats Index (KI) (Kovats 1965) 2865 and 2-methyltriacontane at KI 3065 (Table 1). A density plot of the ln transformed ratios of the peak areas at KI 2865 and KI 3065 [R65 = ln (KI 2865 peak area/KI 3065 peak area)] with an Epanechakov kernel density estimator (Silverman 1986) (Fig. 2) resulted in unimodal distributions for A. suspensa and C. capitata having well separated modes and a small amount of overlap in the distribution tails. No consistent quantitative differences in CHC pattern were evident between feral Puerto Rico and Florida A. suspensa, nor between Hawaii and Guatamala C. capitata. Likewise, the laboratory colonies of C. capitata exhibited R65 values within the range of variation seen in wild collections. The R65 values for feral Florida A. suspensa are consistent with those reported previously (Carlson & Yocom 1986); however, the A. suspensa colony maintained for many years by USDA- ARS, Gainesville, as well as the extant colony at FDACS, Gainesville, exhibited R65

TABLE 1. UNIVARIATE STATISTICS FOR THE LN OF THE RATIO OF THE PEAK AREAS AT KI 2865 AND KI 3065 (R65), FOR THIRD INSTAR LARVAE OF A. SUSPENSA AND C. CAPITATA.

A. suspensa C. capitata N 159 99 Mean 3.45 -0.05 S. D. 2.80 0.15 Minimum 1.43 -3.50 Maximum 4.54 2.14 234 Florida Entomologist 77(2) June, 1994

Fig. 1. Capillary gas chromatograms of hydrocarbons extracted from individual third instar larvae of tephritid fruit flies: a. A. suspensa, b. C. capitata. values substantially lower than those seen in wild collections. This was confirmed by reexamination of CHC patterns from a later sampling (Sutton & Carlson 1993) of the USDA colony. The reduced R65 values in the USDA colony are the consequence of an as yet unidentified eluant of KI 3060 that co-elutes with the KI 3065 peak under the chromatographic conditions used in the previous studies. This peak is also present in wild A. suspensa from Florida and Puerto Rico but in much reduced quantities compared to those seen in individuals from the USDA colony. The origin of the FDACS colony is unclear, but it probably originated from the USDA colony. To avoid this bias, only wild A. suspensa or laboratory colonies recently established from wild individuals were used for comparison with C. capitata. There was no evidence that CHC patterns are correlated with host fruits for either species. Sutton & Steck: Discrimination of Fruit Fly Larvae 235

Fig. 2. Density plot using a kernel density estimator with Epanechakov kernel for the ln transformed ratio of the areas of the 28-and 30-carbon backbone 2-mono- methylalkane peaks (R65) for A. suspensa and C. capitata. Cross-hatching indicates the region of possible overlap.

The specification of a decision criterion and reliability estimates for classification errors based upon R65 values for third instar larvae of A. suspensa and C. capitata depends upon the accurate reconstruction of the distribution tails for each species and the degree of overlap between them. Our sample distributions exhibited a small ≤ ≤ degree of overlap in the region of 1.10 R65 2.30. Given the economic costs of misidentification, it is safer to assume that even this sampling may be insufficient to fully describe population distributions in the range of possible overlap. A linear discriminant model resulted in jack-knifed classification errors of 0.63% and 2.02% for A. suspensa and C. capitata, respectively. However, given that false negative errors are potentially more costly than false positives, a more conservative bias is indicated. The values for A. suspensa drop off sharply for R65 less than 2.30, with only 3% (5 of 159 individuals) below 2.30; a single wild individual had an R65 value less than 2.08. Three percent (3/99 individuals) of the third instar C. capitata larvae exhibited R65 values within the interval (1.10,2.30), with two individuals having R65 values greater than 1.39. Any specific value is somewhat arbitrary given the uncertainties involved, but a classification threshold of about R65 = 2.30 would seem appropriate. 236 Florida Entomologist 77(2) June, 1994

Rare individuals of A. suspensa (3% or so) would be expected to exhibit R65 values below this threshold and would classify as C. capitata. Thus, we must accept the possibility of false positives for larvae having R65 values in the range of 1.10 to 2.30. It should be noted that all third instars analyzed could be correctly identified visually using secondary aspects of the hydrocarbon patterns (Fig. 1). Low R65 values in A. suspensa were due more to variation in the KI 2865 peak than to variation in the KI 3065 peak, i.e., the latter peak remains small with respect to the overall pattern. Similarly, higher R65 values in C. capitata are a consequence of a relative increase in the KI 3065 peak rather than a significant reduction in the KI 2865 peak so that the KI 3065 peak remains large with respect to the overall pattern. Unfortu- nately, such aspects of chromatographic patterns are difficult to quantify, and inter- pretations based upon small peaks can be rendered difficult, if not impossible, by suboptimal chromatographic conditions. Second instar larvae of A. suspensa and C. capitata from laboratory colonies exhibited significant quantitative shifts of the CHC pattern compared to third instars. In both species, second instars show a relative increase in the KI 2865 peak and a reduction in the KI 3065 in comparison to the third instar larvae. A. suspensa from the ± ± FDACS colony exhibited mean R65 values of 4.16 2.09 (n=12) and 2.09 0.88 (n=12) for second and third instars, respectively. No feral second instars of A. suspensa were available for analysis. The R65 values for second and third instar larvae of C. capitata from the lab colony at Waimea were 1.26 ± 0.60 (n=12) and -0.58 ± 0.03 (n=12) respectively. Two feral C. capitata second instars showed corresponding R65 values of 2.36 and 2.81. It is unclear whether there is a more or less continuous shift in the 2-mono- methylalkane series with age, or if the CHC pattern remains relatively invariant over a larval stage. The sampling of second instars was insufficient to construct a realistic classification model; however, it is clear that use of the third instar model would result in a substantial increase in mis-classified C. capitata second instars. Larvae utilized for analysis were either fresh, fresh-frozen or dried. Alcohol preserved specimens generally did not give clean chromatograms and are not recommended for use in CHC analysis. The CHC patterns were not appreciably altered by larval decomposition or physical damage; hence it is possible to utilize specimens that are not identifiable by other means. Very good extractions were also obtained from larval skins alone, e.g. following removal of internal contents for protein electrophoresis or other analysis. Thus, CHC analysis is compatible with simultaneous analysis of a single specimen by both morphological and biochemical or molecular means. Given the possible economic ramifications of misidentification and the ease with which the analysis of a critical individual may be botched, it is imperative that alternative identification systems be available for backup and/or verification. This is particularly important in the case of larvae having R65 values in the region of possible overlap between the two taxa. We found use of 28-200 mesh activated silica gel to be a particularly effective method for drying larvae in the field. While this procedure precludes the use of the larvae for analyses such as protein electrophoresis, solvent extraction of specimens dried in silica gel was significantly enhanced in comparison to fresh or frozen material. Possibly there are highly polar or hydrophilic components in the larval cuticle that shield the hydrocarbons from the non-polar solvent used for extraction, but are removed by adsorption onto the silica gel. Implementation of CHC analysis for the discrimination of larvae of A. suspensa and C. capitata will require further development. No attempt was made in this study to optimize either the larval extraction procedure or the gas chromatography parameters to minimize analysis time and cost. Further sampling will be required to construct a reliable discriminant model for second instar larvae. In addition, the potential exists to adapt and/or extend CHC analysis to the identification of other economically important species, especially Bactrocera, Dacus and other Anastrepha species. Mitchell & Tumlinson: Response to Pheromone 237

ACKNOWLEDGMENT

We gratefully acknowledge the assistance of several colleagues in providing specimens. These include S. Bloem, H. Chang, M. J. Hayes, M. Hennessey P. J. Landolt, L. Stange, and R. Vargas. We also thank D.A. Carlson and S.R. Yocom for reviewing this manuscript.

REFERENCES CITED

CARLSON, D. A., AND S. R. YOCOM. 1986. Cuticular hydrocarbons from six species of fruit flies. Arch. Ins. Biochem. Physiol. 3: 397-412. CLARK, R. A., AND H. V. WEEMS, JR. 1989. Detection, quarantine, and eradication of fruit flies invading Florida. Proc. Florida State Hort. Soc. 102: 159-164. HARDY, D. E. 1949. Studies in Hawaiian fruit flies. Proc. Entomol. Soc. Washington 51: 181-205. HEPPNER, J. B. 1984. Larvae of fruit flies. I. Anastrepha ludens (Mexican fruit fly) and Anastrepha suspensa (Caribbean fruit fly) (Diptera: Tephritidae). Ento- mol. Circ. 260. Florida Dept. Agric. & Cons. Serv. KOVATS, E. 1965. Gas chromatographic characterization of organic substances in the retention index system. Adv. Chromatogr. 1: 229-247. PHILLIPS, V. T. 1946. The biology and identification of trypetid larvae (Diptera: Try- petidae). American Entomol. Soc. Mem. 12, 161 pp. RIHERD, C. 1993. Citrus production areas maintained free of Caribbean Fruit Fly for export certification, pp. 407-413 in M. Aluja and P. Liedo [eds], Fruit Flies: Bi- ology and Management. NY: Springer-Verlag. SILVERMAN, B. W. 1986. Density Estimation for Statistics and Data Analysis. Lon- don: Chapman and Hall, 175 pp. STECK, G. J., L. E. CARROLL, H. CELEDONIO-HURTADO, AND J. GUILLEN-AQUILAR. 1990. Methods for identification of Anastrepha larvae (Diptera: Tephritidae), and key to 13 species. Proc. Entomol. Soc. Washington 92: 333-346. SUTTON, B. D., AND D. A. CARLSON. 1993. Interspecific variation in tephritid fruit fly larval surface hydrocarbons. Arch. Insect Biochem. Physiol. 23: 53-65.

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Mitchell & Tumlinson: Response to Pheromone 237

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RESPONSE OF SPODOPTERA EXIGUA AND S. ERIDANIA (LEPIDOPTERA: NOCTUIDAE) MALES TO SYNTHETIC PHEROMONE AND S. EXIGUA FEMALES1

EVERETT R. MITCHELL AND JAMES H. TUMLINSON Insect Attractants, Behavior, and Basic Biology Research Laboratory, Agricultural Research Service U.S. Department of Agriculture, Gainesville, FL 32604

ABSTRACT

The seasonal occurrence of the beet armyworm, Spodoptera exigua (Hübner), and the southern armyworm, Spodoptera eridania (Cramer), was monitored over a 2-  year period using International Pheromones, Ltd. Moth Traps baited with rubber septa impregnated with pheromone blends identiﬁed from conspeciﬁc females and also with pheromone emitted by live female beet armyworm moths. The pheromone blend for beet armyworm was (Z)-9-tetradecen-1-ol (2.5%), (Z,E)-9,12-tetradecadien-

1This article reports the results of research only. Mention of a proprietary product does not constitute an endorsement or the recommendation of its use by USDA. This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.uﬂ.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

238 Florida Entomologist 77(2) June, 1994

1-ol acetate (87.2%), and (Z)-11-hexadecen-1-ol acetate (10.3%). The pheromone blend used for southern armyworm was (Z)-9-tetradecen-1-ol acetate (55.78%), (Z,E)- 9,12-tetradecadien-1-ol acetate (21.23%), (Z,E)-9,11-tetradecadien-1-ol acetate (8.67%), and (Z)-11-hexadecen-1-ol acetate (14.32%). Although subtropical in habit, both species apparently can survive in North-Central Florida even when temperatures drop below freezing for several days. The response of male southern armyworm moths to traps baited with beet armyworm females was remarkably similar to that recorded for southern armyworm males to their synthetic pheromone. This response was consistent over two years. Key Words: Cross sex attraction, southern armyworm, beet armyworm, (Z)-9-tetradecen-1-ol, (Z,E)-9,12-tetradecadien-1-ol acetate, (Z,E)-9,11-tetradecadien-1-ol acetate, (Z)-11-hexadecen-1-ol acetate.

RESUMEN

La dinámica poblacional del trozador de la remolacha, Spodoptera exigua (Hüb- ner), y del trozador sureño Spodoptera eridania (Cramer), fué estudiada durante 2 años, al usar trampas con feromonas de polillas de la compañía International Pher- omones. El cebo utilizado consistió en un tabique de caucho impregnado con mezclas de feromonas de hembras conespeciﬁcas y con feromonas provenientes de hembras de la polilla de la remolacha. La mezcla de feromona para la polilla de la remolacha fué (Z)-9-tetradecen-1-ol (2.5%), (Z,E)-9,12-tetradecadien-1-ol acetato (87.2%), y (Z)-11-hexadecen-1-ol acetato (10.3%). La mezcla de feromona para el trozador S. eridania fué (Z)-9-tetradecen-1-ol acetato (55.78%), (Z,E)-9,12-tetradecadien-1-ol acetato (21.23%), (Z,E)-9,11-tetradecadien-1-ol acetato (8.67%), y (Z)-11-hexadecen-1-ol acetato (14.32%). Aunque ambas especies son subtropicales, ellas pueden sobrevivir en la parte central y norte de Florida, aún en aquellos días en que la temperatura es bajo zero. La respuesta de las polillas del trozador sureño a las trampas con hembras de polilla de la remolacha fué muy similar a aquellas trampas con feromona sintetica de su misma especie. Esta respuesta fué consistente durante dos años.

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Members of the genus Spodoptera are distributed throughout the world and include a number of species that cause significant economic damage to agricultural crops. The beet armyworm (BAW), S. exigua (Hübner), and southern armyworm (SAW), S. eridania (Cramer), are two of three major North American pests in this genus. Both the BAW and SAW are present in much of Florida throughout the year where they often cause considerable economic damage on commercial vegetable crops. In recent years, the BAW also has become a threat to cotton production in the southeastern U.S. The SAW also has demonstrated a propensity to broaden its host range and include sunflower, often defoliating plants to a degree that yields are greatly reduced (Mitchell 1984). Growers in North-Central Florida are experimenting with alternate crops such as cotton, sunflower, and vegetables to replace declining acreages of tobacco and field corn. Not only are growers planting replacement crops, but they also are investigating ways of intensifying production using double-cropping practices. For example, some growers are following spring planted corn grown for silage with cotton planted in mid-summer; still others are following winter vegetables, such as cabbage, with plantings of sorghum or corn in late spring or early summer. The changing cultural practices have heightened grower awareness of possible shifts in insect pest complexes, especially if plant hosts and suitable habitats become available throughout most of the year. Although subtropical in habit, both the BAW and SAW are potential threats to most of the crops undergoing evaluation by North- Central Florida growers. Therefore, traps baited with synthetic sex pheromone and

Mitchell & Tumlinson: Response to Pheromone 239 live BAW female moths were used to monitor the seasonal occurrence of BAW and SAW in this area over a 2-year period beginning in fall 1988. The synthetic pheromone blend used to monitor the BAW was identified by Tum- linson et al. (1990). They detected and identified five compounds from the volatiles emitted by virgin BAW females: (Z)-9-tetradecen-1-ol acetate; (Z)-9-tetradecen-1-ol; (Z,E)-9,12-tetradecadien-1-ol acetate; (Z,Z)-9,12-tetradecadien-1-ol acetate; and (Z)- 11-hexadecen-1-ol acetate in a ratio of 47.9:4.0:40.2:6.5:1.7, respectively. Field tests of these components in various formulations on rubber septa using bucket traps indicated that a blend of (Z)-9-tetradecenol, (Z,E)-9,12-tetradecadien-1-ol acetate, and (Z)-11-hexadecen-1-ol acetate was most effective for trapping BAW males and that (Z)-9-tetradecen-1-ol acetate actually decreased trap captures. Blends that did not contain both (Z)-9-tetradecen-1-ol and (Z,E)-9,12-tetradecadien-1-ol acetate were ineffective as trap baits (Tumlinson et al. 1990). Analysis of sex pheromone glands and volatile pheromone components collected from calling female SAW indicated that a number of 14-carbon mono- and di-unsaturated acetates and a mono-unsaturated 16-carbon acetate were produced and released (Teal et al. 1985). In the same study, the results of field trapping experiments indicated that a blend of (Z)-9-tetradecen-1-ol acetate (Z9-14:AC), (Z,E)-9,12-tetradecadien-1-ol acetate (Z9,E12-14:AC), (Z,Z)-9,12-tetradecadien-1-ol acetate (Z9,Z12-14:AC), (Z,E)-9,11-tetradecadien-1-ol acetate (Z9,E11-14:AC), and (Z)-11- hexadecen-1-ol acetate (Z11-16:AC) was an effective lure for SAW males capturing significantly greater numbers of moths than did female-baited traps. Of these five components, only Z9,Z12-14:AC did not appear to affect trap capture significantly. Gland extracts also indicated the presence of (Z)-9-tetradecen-1-ol (Z9-14:OH), but this compound was not found in volatiles emitted by females (Teal et al. 1985).

MATERIALS AND METHODS

The study was conducted over a period of 34 months from October 1988 through  June 1991. International Pheromones, Ltd. Moth Traps (i.e., bucket traps), of the type previously used in field evaluations of the pheromone components emitted by calling virgin BAW female moths (Tumlinson et al. 1990), were set out in northwest Alachua County, Florida. Two traps were placed about 1 m above the ground on metal poles at each of three different locations adjacent to cultivated (field corn) and fallowed fields. The distance between traps at each location was about 100 m, and the distance between locations was about 1 km. One of the two traps at each location was baited with four virgin BAW females (2-3 days old). The second trap was baited with a 5 x 9 mm rubber septum (A. H. Thomas Co.) treated with a 100 µl hexane solution containing 200 µg of a pheromone blend previously identified from BAW females: (Z)-9-tetradecen-1-ol (2.5%), (Z,E)-9,12-tetradecadien-1-ol acetate (87.2%), and (Z)-11-hexadecen-1-ol acetate (10.3%) (Tumlinson et al. 1990). In June 1989, a third bucket trap baited with synthetic SAW pheromone was added to each location and positioned as described for the traps containing the BAW females and BAW septum. The SAW septum was treated with a 100 µl hexane solution containing 200 µg of a pheromone blend previously identified from SAW females: (Z)-9-tetradecen-1-ol acetate (55.78%), (Z,E)-9,12-tetradecadien-1-ol acetate (21.23%), (Z,E)-9,11-tetradecadien-1-ol acetate (8.67%), and (Z)-11-hexadecen-1-ol acetate (14.32%) (Teal et al. 1985). No SAW females were available for comparative studies with the synthetic pheromone blend. Septa were Soxhlet-extracted with methylene chloride for 24 h and air dried before loading. The percentage of each component in the pheromone blend loaded onto septa was calculated on the basis of its relative volatility. This was determined from retention indices on liquid crystal capillary GC columns (Heath & Tumlinson 1986)

240 Florida Entomologist 77(2) June, 1994 by a method developed to predict release ratios of components of a blend from rubber septa (Heath et al. 1986).  A Vapona insecticide strip (Vaportape II, Hercon Environmental Company, Emigsville, PA) was placed inside the buckets as a killing agent for trapped moths. The traps were checked 1-3 times per week depending upon the season. During winter (December-February), the traps were checked and rotated between positions weekly. The rest of the year, the traps were checked and rotated on monday, wednes- day, and friday of each week. Bait females were replaced at each inspection; the bait septa and Vaportape II were replaced every two weeks.

RESULTS AND DISCUSSION

Seasonal Response to Pheromone

Captures of BAW in traps baited with rubber septa lures impregnated with the 3- component pheromone blend and virgin BAW females are presented in Fig. 1. Moth captures were greatest during summer and fall both years. Trap capture peaks and valleys for both pheromone and female baits were remarkably similar with the synthetic lure generally capturing more males during most periods. A total of 3,328 BAW males were captured in traps baited with BAW females in 1988-89 and 5,817 were captured in 1989-90. In the traps baited with septa impregnated with pheromone, 7,383 males were captured in 1988-89 and 9,354 were captured in 1989-90. Captures of SAW in traps baited with rubber septa impregnated with synthetic pheromone are shown in Fig. 2. Moth captures were greatest during summer and fall as for the BAW. A total of 2,611 SAW males were captured during 1989-1990 and 1,558 were captured during 1990-1991. The apparent growth in populations of both BAW and SAW, as indicated by the number of moths captured from July through October, was comparatively more uniform during 1990. Although the number of BAW and SAW males captured in pheromone-baited traps during this period were similar to the numbers captured during the same period in 1989, BAW and SAW populations appeared to start increasing about two weeks later in 1990 than in 1989. The steady growth of BAW and SAW moth captures in fall of 1990 likely was due to weather conditions that were conducive to growth of wild hosts common to this area of Florida, especially Amaranthus spp. Based upon weather data compiled by the Agronomy Department, IFAS, University of Florida, Gainesville, drought conditions prevailed throughout Florida from January 1989 through June 1990. During this period, the monthly rainfall deﬁcit, i.e., deviation from expected rainfall, averaged -14.09 cm with a range of -28.21 cm in May 1989 to -0.86 cm in June 1990. The drought was broken in July 1990 and followed by near normal levels of rainfall through November 1990. December 1990 was very dry; the rainfall deﬁcit was -5.38 cm below the normal rainfall level of 8.10 cm for the month. Numbers of BAW and SAW captured in spring and early summer 1991 tended to be lower than numbers captured in the same period of 1990. Again, this probably was due to the level of rainfall received. Unlike spring 1990, which was unusually dry, 1991 was marked by abnormally high levels of rainfall. From January through July 1991, rainfall averaged 17.98 cm above normal, ranging from +0.33 cm in February to +26.41 cm in July. The torrential rains that often occurred during this period possibly helped keep the armyworm populations low, by destroying young larvae soon after they hatched, and also by increasing the level of natural control via disease organisms. Both BAW and SAW are subtropical species that cannot diapause and thus are unable to survive during extended periods of near- or sub-freezing temperatures. The species' overwintering range appears to be governed by the occurrence of frost

Mitchell & Tumlinson: Response to Pheromone 241

Fig. 1. Weekly captures of male beet armyworm moths in bucket traps baited with rubber septa lures treated with synthetic sex pheromone simulating conspeciﬁc females and beet armyworm females. Numbers above months are reference dates for beginning of weekly periods. October 1988-September 1990. Alachua County, Florida.

242 Florida Entomologist 77(2) June, 1994

Fig. 2. Weekly captures of male southern armyworm moths in bucket traps baited with rubber septa lures treated with synthetic sex pheromone simulating conspeciﬁc females and with natural sex pheromone emitted by beet armyworm females. Num- bers above months are reference dates for beginning of weekly periods. July 1989- June 1991. Alachua County, Florida.

Mitchell & Tumlinson: Response to Pheromone 243 that kills host plants and by prolonged temperatures below about 10°C. Thus, the BAW and SAW are very similar in habit, overwintering by continuous generations in subtropical areas of Florida and Texas, and in a few areas along the Gulf Coast (Mitchell 1979). Both species appear to disperse from these areas into more temperate zones of the United States each spring and summer. The data on moth captures for the winter of 1989-1990 and 1990-1991 present paradoxical situations relative to survival of BAW and SAW in North-Central Flor- ida. Three days of continuous sub-freezing temperatures were recorded in this area during late December 1989, with snow and ice recorded on the 23rd and 24th. Depar- tures from the normal maximum, minimum and average temperature for December were -23o, -23.7o, and -23.4o C., respectively. The effect of the unusually cold temperatures was reﬂected in zero moth captures in traps baited with BAW females and synthetic SAW pheromone, respectively, during the last two weeks of December 1989 and the ﬁrst week of January 1990. Never- theless, a few BAW and SAW males were captured early in January 1990, and captures of both species continued into spring. These results suggest that the BAW and SAW possibly survived winter in North-Central Florida even under the rather extreme conditions of below freezing temperatures. Of course, the possibility that the insects captured during this period were migrants cannot be discounted completely. In contrast to winter 1989-1990, the winter of 1990-1991 was extremely mild, allowing continuous growth of herbaceous species that normally would have been killed by frost. These conditions probably allowed for reproduction by BAW and SAW later in fall 1990; hence, accounting for possible local emergence of the moths captured during December 1990-January 1991. Comparative data on capture of BAW in traps baited with synthetic pheromone in the same period are not available because these traps were discontinued in October 1990.

Cross Sex Attraction

Mitchell & Doolittle (1976) reported cross sex attraction between BAW and SAW. This intriguing relationship is demonstrated also in these studies (Figs. 2 and 3).  Mitchell and Doolittle used females of each species in Pherocon 1C sticky traps and about 13% of the moths captured in traps baited with BAW females were SAW males. Conversely, about 8% of the males captured in traps baited with SAW females were BAW males. In the present study, a total of 8,420 BAW males were captured in traps baited with BAW females from July 1989 through June 1991 (2,742 in 1989-1990 and 5,678 in 1990-1991). A total of 2,019 SAW males also were captured in the same traps (1,253 in 1989-1990 and 766 in 1990-1991). A total of 4,169 SAW males were captured in traps baited with lures treated with synthetic SAW sex pheromone. However, no BAW males were captured in any of these traps over the course of the two-year survey. Interestingly, the percentage of SAW males captured in traps baited with BAW females equaled 48.4% of the number of SAW males captured in traps baited with synthetic SAW sex pheromone. The pattern of capture of SAW males in traps baited with BAW females paralleled closely the capture of SAW in traps baited with synthetic SAW sex pheromone (Fig. 2). The results of regression analysis of the number of SAW males captured in traps baited with BAW females versus the numbers captured in traps baited with synthetic SAW pheromone are presented in Fig. 3. These results suggest that cross sex attraction of SAW males to BAW females was consistent and independent of possible seasonal effects that may have been caused by positioning the traps close to host plants. Moreover, the capture of SAW males in traps baited with BAW females was as good an indicator of seasonal occurrence and population peaks of SAW (Fig. 2), as was the synthetic pheromone blend identiﬁed from conspeciﬁc females.

244 Florida Entomologist 77(2) June, 1994

Fig. 3. Relationship between the number of male southern armyworm moths captured in traps baited with beet armyworm females and numbers captured in bucket traps baited with synthetic sex pheromone simulating pheromone produced by southern armyworm females.

Mitchell & Tumlinson: Response to Pheromone 245

The reason for attraction of SAW males to BAW females is indeed puzzling. How- ever, the chemical composition of the sex pheromone blend volatized by the two species may partially explain the unusual cross attraction reported here. The BAW and SAW have four chemicals in common in their pheromone blend (Table 1). Of these, (Z)-9-tetradecen-1-ol acetate is the major component volatized by females of both species; it is a critical component for attraction of SAW males (Teal et al. 1985) but not for BAW (Tumlinson et al. 1990).The role of (Z)-9-tetradecen-1-ol acetate in the sexual behavior of BAW is unknown; but, addition of this compound to the attractive pheromone blend (Table 1) actually decreases capture of BAW males. Interestingly, (Z)-9-tetradecen-1-ol was not found among the chemicals volatized by calling SAW females (Teal et al. 1985), but it was found in gland extracts from SAW females. It is possible, of course, that (Z)-9-tetradecen-1-ol was emitted by SAW females but escaped detection because it occurs in such minute quantities. This component is essential to attraction of BAW males (Tumlinson et al. 1990), and it is volatized by BAW females. When Teal et al. (1985) added (Z)-9-tetradecen-1-ol to the SAW pheromone blend, trap captures of SAW were not affected. In an earlier study, Mitchell (1976) found no evidence of cross mating among these species in the laboratory. Thus, it is unlikely that SAW males would mate with BAW females in nature even when attracted to them. Perhaps (Z)-9-tetradecen-1-ol acts as a reproductive isolating mechanism for BAW and SAW during close range courtship encounters. The design of the bucket trap used in this study is such that the insects captured must come within a few centimeters of the lure. Observations on the response of other noctuid species, namely S. frugiperda (J. E. Smith) and Anticarsia gemmatalis Hübner, to synthetic sex pheromone lures in bucket traps indicated that the male moths became very excited when in close proximity to the bait source. Generally, the frenzied moths tumbled into the bucket receptacle after 'bumping' into the wall of the open funnel immediately beneath the lure. Therefore, it is probable that chemicals other than those previously identiﬁed from females of the BAW and SAW are responsible for their reproductive isolation in nature.

TABLE 1. VOLATILE PHEROMONE COMPONENTS EMITTED BY FEMALES OF SPODOPTERA EXIGUA (BAW) AND S. ERIDANIA (SAW). A (+) INDICATES THE COMPONENT IS NECESSARY IN THE PHEROMONE BLEND FOR CAPTURING CONSPECIFIC MALES IN NUMBERS = OR > THAN THE NUMBERS CAPTURED IN THE SAME TYPE OF TRAP BAITED WITH VIRGIN FEMALES OF THE SAME SPECIES. ADDI- TION OF THE COMPONENT INDICATED BY (*) TO THE PHEROMONE BLEND RE- SULTED IN CAPTURE OF BAW MALES. ADDITION OF THE COMPONENT INDICATED BY (!) TO THE PHEROMONE BLEND DIMINISHED CAPTURE OF BAW MALES. Chemical1 BAW2 SAW3 Z9,E12-14:AC (+) (40.2)4 (+) (16.9)4 Z9,Z12-14:AC (6.50) (15.2) Z9,E11-14:AC (+) (4.60) Z9-14:OH (+) (4.00) (*) Z9-14:AC (!) (47.9) (+) (60.6) Z11-16:AC (+) (1.70) (+) (2.70)

1 See text for explanation of chemical abbreviations. 2Tumlinson et al. 1990. 3Teal et al. 1985. 4Numbers in () are release ratios [%] of pheromone components volatized by calling females. Load ratios used to achieve the desired release ratios when applied on rubber septa used as lures in bucket traps for SAW were: Z9,E12-14:AC, 21.23%; Z9,E11-14:AC, 8.67%; Z9-14:AC, 55.78%; and Z11-16:AC, 14.32%. 246 Florida Entomologist 77(2) June, 1994

CONCLUSIONS

Synthetic pheromone lures for the BAW were as reliable an indicator of seasonal occurrence as were virgin BAW females. Although trap catches with the synthetic blend are well-correlated with live females, this does not necessarily mean that the catches with either of these lures is well-correlated with actual population densities in any given field. Indeed, the senior author often has recorded large captures of these species as well as others including S. frugiperda (J. E. Smith), Heliothis virescens (Fabricius), and Helicoverpa zea (Boddie), in traps baited with synthetic pheromone; but the trap catches usually did not reflect population densities based upon actual egg, larval, or adult counts in crops such as cotton, corn, and sorghum (unpublished). Although pheromone release rates from septa are influenced by temperature, there was no indication that the cooler temperatures of winter and spring or the hot, humid conditions of summer in North-Central Florida had any more influence on capture of BAW in traps baited with synthetic pheromone than captures in similar traps baited with virgin BAW females. Indeed, the economy of labor, ease of handling, and consistency of delivering a precise quantity of a pheromone at a relatively modest cost, compared with rearing and maintaining live insect baits and the confi- dence that the results obtained with synthetic lures are as meaningful as those obtained with live insects, warrants the continued use of synthetic pheromone for long- term monitoring of BAW. Also, the synthetic sex pheromone lure used for the SAW appeared to provide a realistic picture of the seasonal occurrence and shifts in population numbers of this pest. Although SAW female baits were not available for comparison with the synthetic lure, it is not unreasonable to assume that a comparative study would have yielded results similar to those recorded for the BAW. Captures of SAW males in traps baited with BAW females presented as accurate an assessment of its seasonal occurrence as was defined by captures of SAW males in traps baited with synthetic SAW pheromone. As desirable as such a dual monitoring scheme may be, the use of BAW females to monitor SAW populations is unrealistic. As demonstrated here, synthetic BAW pheromone, not BAW females, is the bait of choice for monitoring this pest. Unfortunately, SAW males are not captured in traps baited with synthetic BAW pheromone.

ACKNOWLEDGMENT

We gratefully acknowledge the technical assistance of P. Brennan for preparing the bait septa, W. Copeland for collecting the trap data, and J. Leach for preparation of the graphic material.

REFERENCES CITED

HEATH, R. R., AND J. H. TUMLINSON. 1986. Correlation of retention times on a liquid crystal capillary column with reported vapor pressures and half-lives of compounds used in pheromone formulations. J. Chem. Ecol. 12: 2081-2088. HEATH, R. R., P. E. A. TEAL, J. H. TUMLINSON, AND L. J. MENGELKOCH. 1986. Predic- tion of release of multicomponent pheromone blends from rubber septa. J. Chem. Ecol. 12: 2133-2143. MITCHELL, E. R. 1976. Mating speciﬁcity in Spodoptera spp. Florida Entomol. 59: 416. MITCHELL, E. R. 1979. Migration by Spodoptera exigua and S. frugiperda, North American style, pp. 386-393 in R. L. Rabb and G. G. Kennedy [eds.] Movement of highly mobile insects: concepts and methodology in research. University Graphics, North Carolina State University, Raleigh. Giblin-Davis et al.: Trapping of Metamasius hemipterus 247

MITCHELL, E. R. 1984. Damage of sunflower by the southern armyworm (Lepi- doptera: Noctuidae). Florida Entomol. 67: 273-277. MITCHELL, E. R., AND R. E. DOOLITTLE. 1976. Sex pheromones of Spodoptera exigua, S. eridania, and S. frugiperda: Bioassay for field activity. J. Econ. Entomol. 69: 324-326. TEAL, P. E. A., E. R. MITCHELL, H. H. TUMLINSON, R. R. HEATH, AND H. SUGIE. 1985. Identification of volatile sex pheromone components released by the southern armyworm, Spodoptera eridania (Cramer). J. Chem. Ecol. 6: 717-725. TUMLINSON, J. H., E. R. MITCHELL, AND H. -S. YU. 1990. Analysis and field evaluation of volatile blend emitted by calling virgin females of beet armyworm, Spodoptera exigua (Hübner). J. Chem. Ecol. 16: 3411-3423.

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Giblin-Davis et al.: Trapping of Metamasius hemipterus 247

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LETHAL PITFALL TRAP FOR EVALUATION OF SEMIOCHEMICAL-MEDIATED ATTRACTION OF METAMASIUS HEMIPTERUS SERICEUS (COLEOPTERA: CURCULIONIDAE)

1 ROBIN M. GIBLIN-DAVIS,1 JORGE E. PEÑA,2 AND RITA E. DUNCAN 1Fort Lauderdale Research and Education Center University of Florida, IFAS 3205 College Avenue, Fort Lauderdale, FL 33314

2Tropical Research and Education Center University of Florida, IFAS 18905 S. W. 280 Street, Homestead, FL 33031

ABSTRACT

A lethal pitfall trap was developed for evaluating field response of adults of the West Indian sugarcane borer, Metamasius hemipterus sericeus (Olivier), to semiochemical attractants. Using this trap, we determined that early (≤ 5 days) volatile fermentation products from 250 g of chopped fresh stem tissue from the cabbage palmetto (Sabal palmetto [Walter]), sugarcane (Saccharum officinarum L.), pineapple (Anana comosus [L.]), or pseudostem from banana (Musa sp.) (fermented under water for 5-7 days) were attractive to M. h. sericeus adults in the field. Weevils were not trapped with fresh banana pseudostem and chopped unfurled leaves of S. palmetto. The addition of 20 males or 20 females to 250 g of chopped S. palmetto stem tissue increased capture of M. h. sericeus over tissue alone. Surveys with the lethal pitfall trap baited with sugarcane tissue demonstrated that M. h. sericeus is established in Dade, Broward, and Palm Beach Counties, Florida. This weevil poses a potential threat to sugarcane and field plantings of certain ornamental palm species, including Hyophorbe verschaffeltii Wendland, Phoenix canariensis Hortorum ex Chabaud, Ptychosperma macarthurii (Wendland), Ravenea rivularis, Roystonea regia (Hum- bolt, Bonpland & Kunth), and Washingtonia robusta Wendland.

RESUMEN

Se evaluó en el campo la atracción de adultos del picudo de la caña podrida, Meta- masius hemipterus sericeus (Oliver) a atrayentes químicos colocados en una trampa letal. Los primeros productos volátiles (< 5 días) emanados de la fermentación de 250 g de tejido fresco picado de palmeto de col (Sabal palmetto [Walter]), caña de azúcar (Saccharum ofﬁcinarum L.), piña (Anana comosus [L.]) o pseudotallo de banano (Musa sp.) (fermentado en agua por 5-7 días), fueron atractivos a los adultos de M. h.

248 Florida Entomologist 77(2) June, 1994 sericeus en el campo. El pseudotallo de banano y las hojas picadas del cogollo de S. palmetto no atrayeron adultos del picudo. La captura de M. h. sericeus aumentó al adicionar 20 machos o 20 hembras a una trampa con 250 g de tallo picado de S. palmeto en comparación con el tallo picado solo. El uso de la trampa letal demostró que M. h. sericeus está establecido en Florida en los condados de Dade, Broward y Palm Beach, Florida y presenta una amenaza potencial a la caña de azúcar y a ciertas especies de palmas ornamentales, tales como Hyophorbe verschaffeltii Wendland, Phoenix canariensis Hortorum ex Chabaud, Ptychosperma macarthurii (Wendland), Ravenia rivularis, Roystonia regia (Humboldt, Bonpland & Kunth), y Washingtonia robusta Wendland.

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The West Indian sugarcane borer, Metamasius hemipterus (L.) is an economically important pest of sugarcane and other tropical plants in the Neotropics (Raigosa 1974, Vaurie 1966). There are three recognized subspecies of this weevil; M. h. hemipterus (L.) is distributed southwards from Puerto Rico through the Lesser Antilles and into most of South America; M. h. sericeus (Olivier) is found in the Greater Anti- lles and Central America south from Nicaragua to western Colombia and Ecuador; while M. h. carbonarius (Chevrolat) occurs from Mexico south to El Salvador and Honduras (Vaurie 1966). Metamasius hemipterus sericeus first became established in Dade Co., Florida in about 1984 (Woodruff & Baranowski 1985). The biology of all three subspecies is very similar. Adult weevils are attracted to, and oviposit in, damaged or stressed sugarcane stalks, banana pseudostems, ripe fruit (i.e. pineapple, mango, papaya), or palm sheaths or stems (Vaurie 1966) where the larvae develop to adults in less than 2 months (Woodruff & Baranowski 1985). Wolcott (1955) reported that the female weevil oviposits in damaged sugarcane stem but that the larvae tunnel into healthy tissue causing it to ferment. Fermenting sugarcane tissue has been used to attract M. hemipterus (Raigosa 1974) and M. anceps (Gyllenhal) (Teran 1968) in the field. Also, a yeast, water, and molasses mixture applied to sugarcane bagasse in single open-ended bamboo cylinder traps was used to attract M. hemipterus in Colombia (Arango & Rizo 1977). In all of these cases, the bait was poisoned with an insecticide to kill the weevils. Six times more M. hemipterus were attracted to sugarcane soaked for 12-24 hrs in water than to sugarcane soaked in 0.1% parathion and placed in traps made of 30-40 cm long cyl- inders of bamboo with a single open end (Raigosa 1974). The use of a lethal trap is critical for evaluation of semiochemicals, especially when pheromones are suspected. The use of insecticides can be very effective for field trapping and study of the chemical ecology of weevils (Oehlschlager et al. 1992). However, the attractancy or repel- lency of the insecticide to the target organism must be evaluated and safety precautions must be followed in trap set-up, placement, and maintenance. A safe and effective trap for evaluating semiochemical attractants for the subspecies of M. hemipterus is needed. Males of many species of weevils produce aggregation pheromones which can be useful for monitoring the movement of pests, mass trapping, or for timing applications of pesticides or biologicals (Weissling et al. 1994). Increasing our knowledge of semiochemical attractants for M. h. sericeus is important because they would be extremely useful for monitoring weevil movement into the southeastern U.S.A. and for developing biorational approaches for their management in Florida. The purpose of this study was twofold: (1) to develop a safe, lethal trap to field-evaluate the attraction of semiochemicals to M. h. sericeus adults and (2) to use this trap to evaluate the attraction of different fermenting plant tissues alone, and with or without conspecifics, to M. h. sericeus adults in the field. During this study we also demonstrated that this weevil is newly established in two counties in southeastern Florida. Symptoms of M. h. sericeus infestations in field-grown palm plantings were observed and are described herein.

Giblin-Davis et al.: Trapping of Metamasius hemipterus 249

MATERIALS AND METHODS

Lethal Trap Design

The lethal pitfall trap that was developed consisted of two translucent white, high density polyethylene (HDPE) plastic tapered containers (Fisher Scientiﬁc Co., Pittsburgh, PA), one 4.9 liters and the other 0.946 liters in size (Fig. 1). Soapy water in the bottom of the larger container acted as the lethal agent. Four 3-mm diam drainage holes were drilled at 90° from each other in the sides of the 4.9-liter bucket, 6 cm from the bottom. The 0.946 liter container, which was used to hold the attractant, had 2 rows of 9 ventilation holes drilled in its sides near the bottom (Fig. 1b). An 8.5-cm diam hole was cut in the lid of this container and a 9.5-cm diam piece of alu- minum screening (1.5-mm square openings) was glued to the inside of the lid to cover this hole. The lid was needed to retain the attractant (250 g of chopped test tissue and/or adult weevils) within the container when it was inverted inside the larger container (Fig. 1b). It was suspended in this container on a vinyl clad steel wire (32 cm in length) which was threaded through 3-mm holes at 180° from each other 1 cm below the top opening in the 4.9-liter bucket and through the bottom (now top) of the inverted bait container. It was suspended concentrically in the mouth of the bucket (Fig. 1a,b). Each assembled trap was buried to within about 2.5 cm of the trap top in the shade of a mature banana plant and ﬁlled with about 600 ml of soapy water (10 g Alconox soap [Alconox, Inc., New York, NY] per liter water). In a preliminary study, this trap design was baited with 250 g of banana (Musa sp.) pseudostem (fermented under water for 5-7 days) and compared with the following six designs: 1) the above design set on the soil surface, 2) a fermented banana pseudostem sandwich trap (Mitchell 1978) with two slices of pseudostem (total about

Fig. 1. Lethal pitfall trap used to capture and kill adults of Metamasius hemipterus sericeus attracted to different fermenting plant tissues and/or conspeciﬁcs: (a) assembled, (b) unassembled.

250 Florida Entomologist 77(2) June, 1994

550 g of tissue) bound together with a rubber band and placed on the ground, 3) a live Rhynchophorus cruentatus (F.) trap (Weissling et al. 1992) placed on the ground with 1.3 kg of fermented banana pseudostem, 4) a lethal R. cruentatus trap (Weissling et al. 1993) set on the ground, with and without a polyvinyl chloride composite lid, baited with 1.3 kg of fermented banana pseudostem, and 5) a 19-liter black plastic bucket set on the ground and baited with 1.3 kg of fermented banana pseudostem. There were two replicates for each of the seven designs and traps were set out in a 15.5 ha, 2 year-old banana plantation, located near Homestead, Dade County, FL. Traps were spaced ≥ 20 m apart in rows of banana plants.

Field Response of Weevils to Fermenting Plant Tissues

Experiment No. 1 was conducted 6-11 January 1993 using the lethal pitfall trap described previously. The plant tissue attractants were 250 g of chopped tissue from the following sources: 1) fresh unfurled leaves of Sabal palmetto (Walter), 2) fresh stems of sugarcane, Saccharum ofﬁcinarum L., 3) fresh pineapple, Anana comosus (L.), 4) fresh pseudostem of banana, Musa sp., 5) 7-10 cm-thick cross-sections of banana pseudostem fermented for 5-7 days under water, and 6) no tissue. Experiment No. 2 was conducted 20-25 January 1993. The treatments were the same as for experiment No. 1 except that both banana pseudostem treatments were dropped and fresh S. palmetto stem tissue was added as the ﬁfth treatment. Traps for both experiments were arranged in a randomized complete block design in the previously described banana plantation. Treatments were spaced about 20 m apart in a row of banana plants (block) and blocks were separated by about 20 m (2 or more rows of banana plants). There were seven blocks for experiment No. 1 and eight for experiment No. 2. Total numbers of adult male and female M. h. sericeus collected per trap for each 5-day trapping period were used for analysis. Data were transformed (x + 0.5)0.5 and a Statistical Analysis System general linear models procedure (SAS Institute 1985) for unbalanced ANOVA was used. Untransformed means are presented. A Waller-Duncan k-ratio t-test (k = 100, P ≤ 0.05) was used for means separation.

Field Response of Weevils to Fermenting Plant Tissues and/or Conspeciﬁcs

The lethal pitfall trap (Fig. 1) was used in the field to test the attraction of weevils to live conspecifics in contact with and without S. palmetto tissue. All traps contained two water-moistened cellulose sponges. Treatments were traps baited with the following: 1) moistened sponges (unbaited control), 2) 20 males of M. h. sericeus, 3) 20 females of M. h. sericeus, 4) 250 g of chopped fresh S. palmetto stem tissue, 5) 250 g of chopped tissue plus 20 male weevils, 6) 250 g of chopped tissue plus 20 female weevils, and 7) 250 g of banana pseudostem fermented as previously described. The experiment was conducted 2-7 December 1992 (4 replicates) and 9-14 December 1992 (4 replicates) after re-randomization of treatments in a different field location. Ex- perimental design and analysis were as previously described.

Survey of Dade, Broward, and Palm Beach Counties in Florida for M. h. sericeus and Symptoms

Lethal traps baited with 250 g of chopped sugarcane were used to survey plantings of sugarcane, banana, or ornamental palms in Broward (3 sites), Dade (10 sites), and Palm Beach (8 sites) counties during March-September 1993. Two to seven survey traps were used in each location. Broward, Dade, and Palm Beach county extension units, personnel of the Florida Department of Agriculture and Consumer Services, Division of Plant Industry, and cooperative growers collaborated with us in Giblin-Davis et al.: Trapping of Metamasius hemipterus 251 locating active M. h. sericeus infestations and candidate sites for survey and observation of symptoms in host plants.

RESULTS AND DISCUSSION

Lethal Trap Design

In the preliminary study, the lethal pitfall trap (Fig. 1) was 2-17 times more effective for trapping of M. h. sericeus than any of the other designs tested. The mean counts from the traps tested were; lethal pitfall trap depicted in Fig. 1 (17 weevils), trap design 1 (2 weevils), trap design 2 (5 weevils), trap design 3 (1 weevil), trap design 4 with pvc lid (9 weevils), trap design 4 without pvc lid (9 weevils), and trap design 5 (1 weevil). The lethal trap is safe, simple, effective, and kills incoming weevils, thus preventing contact with test attractants or pheromone(s). This design was used for all of our other experiments. The trap design, color, and placement will be optimized in future studies when other semiochemical attractants for M. h. sericeus have been identiﬁed.

Field Response of Weevils to Fermenting Plant Tissues

Our study showed that semiochemicals produced by early fermentation (≤ 5 days) of a variety of plant tissues were equally attractive to M. h. sericeus. In experiment No. 1, sugarcane tissue was signiﬁcantly more attractive to females of M. h. sericeus (Table 1) than pineapple and fermented banana pseudostem, which were equally attractive. Unfurled leaves of S. palmetto, fresh banana pseudostem, and the check were equally unattractive (Table 1). Sugarcane and pineapple were equally attractive to males of M. h. sericeus. The other tissues were not signiﬁcantly different from the check (Table 1). The combined sex ratio for all treatments was about 5.5 females to 1.0 male. In experiment No. 2, S. palmetto stem tissue, sugarcane, and pineapple were all equally attractive to males and females of M. h. sericeus. Unfurled leaves of S. palmetto and the check were not attractive (Table 1). The sex ratio in experiment No. 2

TABLE 1. ATTRACTION OF METAMASIUS HEMIPTERUS SERICEUS ADULTS TO LETHAL FIELD TRAPS BAITED WITH DIFFERENT PLANT TISSUES (250 G) IN A BANANA PLANTATION IN DADE CO., FLORIDA, FOR 5 DAYS. No. Weevils per Trap (Mean ± S.E.)1 No. of Treatments Replicates Females Males Experiment No. 1 (6-11 January 1993)

Fresh sugarcane stem (chopped) 7 69.6 ± 23.6a 11.4 ± 3.6a Fresh pineapple (chopped) 6 25.0 ± 15.3b 7.3 ± 4.9ab Fermented banana pseudostem2 (chopped) 7 16.3 ± 6.0b 1.3 ± 0.7bc Unpresented Sabal palmetto leaves (chopped) 7 1.3 ± 0.5c 0.4 ± 0.3bc Fresh banana pseudostem (chopped) 7 0.7 ± 0.3c 0.0 ± 0.0c Empty 7 0.0 ± 0.0c 0.0 ± 0.0c 252 Florida Entomologist 77(2) June, 1994

TABLE 1. (CONTINUED) ATTRACTION OF METAMASIUS HEMIPTERUS SERICEUS ADULTS TO LETHAL FIELD TRAPS BAITED WITH DIFFERENT PLANT TISSUES (250 G) IN A BANANA PLANTATION IN DADE CO., FLORIDA, FOR 5 DAYS. No. Weevils per Trap (Mean ± S.E.)1 No. of Treatments Replicates Females Males Experiment No. 2 (20-25 January 1993)

Fresh Sabal palmetto stem (chopped) 8 3.8 ± 1.5a 25.4 ± 12.2a Fresh sugarcane stem (chopped) 8 2.9 ± 2.3ab 18.9 ± 6.4a Fresh pineapple (chopped) 8 1.3 ± 0.5ab 15.8 ± 7.7a Unpresented Sabal palmetto leaves (chopped) 8 0.0 ± 0.0b 0.3 ± 0.2b Empty 8 0.0 ± 0.0b 0.0 ± 0.0b

1Means followed by the same letter are not signiﬁcantly different according to a Waller-Duncan k-ratio t-test on (x + 0.5)0.5 transformed data (k = 100, P ≤ 0.05). Untransformed means are presented. 2Fermented as 7-10 cm thick cross-sections under water for 5-7 days. was reversed from that in experiment No. 1 with about 7.5 males to 1.0 female. The lack of attractancy of fresh banana pseudostem and unfurled leaves of S. palmetto may have been caused by a lack of moisture in these tissues. This would have affected the quality of fermentation and the quantity of attractive volatiles released. Surprisingly, S. palmetto stem tissue was an excellent source of attractive volatiles even though this palm has not been observed as a host for M. h. sericeus. These data are consistent with observations that M. h. sericeus is attracted to wounded or fermenting tissue of palms, sugarcane, banana, and certain fruits (Vaurie 1966). Cur- rently, we are using gas chromatograph-electroantennograph detection (GC-EAD) procedures to identify semiochemicals attractive to M. h. sericeus which have been collected from air perfusions of fermenting S. palmetto, sugarcane, pineapple, and fermented banana pseudostem. Active chemicals are being evaluated in the ﬁeld using the lethal pitfall trap.

Field Response of Weevils to Fermenting Plant Tissues and/or Conspeciﬁcs

Sabal palmetto tissue plus 20 males and S. palmetto tissue plus 20 females were equally attractive to females and males of M. h. sericeus and more attractive than S. palmetto tissue alone (Table 2). Twenty females or 20 males alone, fermented banana pseudostem alone, and the empty check were all equally not attractive to males and females (Table 2). Although not statistically significant, male M. h. sericeus increased the attractiveness of palm tissue more than females (Table 2). Our data show that semiochemicals emanating from fermenting tissue of S. palmetto are attractive by themselves (Table 2). The feeding activity of males or females on fermenting tissue possibly enhances the release of these volatiles (Table 2). This has been observed when the pine weevil Hylobius abietis (L.) feeds on pine (Tilles et al. 1986). These results differ from what has been observed for the palmetto weevil, Rhynchophorus cruentatus (F.). With this species the combination of caged males and S. palmetto tissue was much more attractive than females and tissue, tissue alone, males alone, females alone, and the empty check, all of which were about equally unattractive (Weissling et al. 1993). A male-produced pheromone, 5-methyl-4-octanol (cruentol), has been identified from R. cruentatus. It has been shown to be ineffective in field traps when released alone but strongly synergizes the weak attraction of volatiles from fermenting plant tissue (Giblin-Davis et al. 1994, Weissling et al. 1994). Giblin-Davis et al.: Trapping of Metamasius hemipterus 253

TABLE 2. ATTRACTION OF METAMASIUS HEMIPTERUS SERICEUS ADULTS TO LETHAL FIELD TRAPS BAITED WITH FRESH SABAL PALMETTO OR FERMENTED BA- NANA PSEUDOSTEM TISSUES (250 G) AND/OR LIVE CONSPECIFIC WEEVILS, OR LEFT UNBAITED IN A BANANA PLANTATION IN DADE CO., FLORIDA FOR 5 DAYS (2-7 AND 9-14 DECEMBER 1992). No. Weevils per Trap (Mean ± S.E.)1 No. of Treatments Replicates Females Males Fresh S. palmetto stem + 20 (( M. hemipterus 8 36.1 ± 8.6a 24.6 ± 5.8a Fresh S. palmetto stem + 20 && M. hemipterus 8 30.8 ± 5.4a 17.6 ± 3.9a Fresh S. palmetto stem 8 18.1 ± 4.9b 6.0 ± 2.3b Fermented banana pseudostem2 7 3.7 ± 1.2c 1.0 ± 0.8c 20 (( M. hemipterus 8 0.4 ± 0.2c 0.0 ± 0.0c Empty 8 0.0 ± 0.0c 0.3 ± 0.3c 20 && M. hemipterus 8 0.0 ± 0.0c 0.1 ± 0.1c

Recently, we collected volatiles from males and females of M. h. sericeus and recovered eight male-produced chemicals (four GC-EAD-active alcohols and four GC-EAD-active ketones) (Perez et al. 1994). Using the lethal pitfall trap described herein (Fig. 1), the four alcohols combined (3-pentanol, 2-methyl-4-heptanol, 2-methyl-4-octanol, and 4-methyl-5-nonanol), each released at 3 mg/day, were as attractive as 250 g of sugarcane. When combined, these two treatments showed addi- tive, not synergistic, attraction (Perez et al. 1994). The four GC-EAD-active ketones of M. h. sericeus males reduced weevil attraction in the ﬁeld when released with the GC-EAD-active alcohols plus sugarcane suggesting a role as “spacing” pheromones (Perez et al. 1994). The lack of a signiﬁcant difference in attraction between treatments of males plus palm tissue and females plus palm tissue in the present study is not clear. Densities of 20 males of M. h. sericeus may have been too high resulting in a release of the four GC-EAD-active ketones negating any response to attractive male-produced semiochemicals.

Survey of Dade, Broward, and Palm Beach Counties in Florida for M. h. sericeus and Symptoms

Metamasius hemipterus sericeus were trapped near and in banana plantings, sugarcane fields, and fields of ornamental palms in southern Dade County, and Pa- hokee and Belle Glade, Palm Beach County. We also observed larval infestations for the first time in damaged sugarcane at the Everglades Research and Education Cen- ter in Belle Glade (1 February 1994). In central and western Broward County, M. h. sericeus adults were trapped near and in small banana plantings and fields of ornamental palms. Larval infestations of M. h. sericeus were noted in palms during 1991-1993 in Florida as follows: 1) forty healthy (Dade Co.) and about 20 wounded (Dade and Bro- ward Co.) 3 to 4-year-old, field grown spindle palms, Hyophorbe verschaffeltii Wend- land; 2) four 3 to 4-year-old healthy Canary Island date palms, Phoenix canariensis Hortorum ex Chabaud in Dade Co.; 3) three healthy MacArthur palms, Ptycho- sperma macarthurii (Wendland) in Dade Co.; 4) more than one hundred 3 to 254 Florida Entomologist 77(2) June, 1994

4-year-old healthy field grown Majesty palms, Ravenea rivularis in Palm Beach Co.; 5) six 3 to 4-year-old healthy field grown royal palms, Roystonea regia (Humbolt, Bonpland & Kunth) in Broward and Palm Beach Co.; and 6) two 3 to 4-year-old healthy Washington fan palms, Washingtonia robusta Wendland in Dade Co. Metamasius hemipterus sericeus larval tunneling appeared to start in petioles or wounds in the petioles, crown, or stem and then extends into healthy stem tissue. Observed symptoms included: 1) the appearance of a dark amber-colored gummy ex- udate which issued from openings to the surface from larval galleries in the stem and petioles of the crown region or sometimes in the stem near or amongst exposed roots (R. regia); and 2) open 1.0-1.5 cm diam larval galleries in the leaves, petioles, and stem. Signs included abandoned cocoons made of stem or petiole fibers, and/or adult, larval, and pupal weevils at the base of petioles and in galleries in the stem. Overall symptoms were a lethal wilt with general chlorosis and premature leaf death which was noticed in all of the P. macarthurii, and in several H. verschaffeltii and R. rivularis that were examined. Eventually, the crown was completely destroyed and col- lapsed because of larval tunneling. These symptoms are very similar to those caused in a number of palm species by the cane weevil borer, Rhabdoscelus obscurus (Bois- duval) in Australia (Halfpapp and Storey 1991). Because the smooth, columnar trunks of H. verschaffeltii, R. rivularis, and R. regia are an important part of their aesthetic appeal, even slight damage by M. h. sericeus is of economic importance. The W. robusta that we examined were booted (with old petioles attached) and asymptomatic for M. h. sericeus damage until the boots were removed near the crown revealing typical larval weevil damage. Booted species of palms would have a higher threshold for aesthetic trunk damage but high densities of undetected M. h. sericeus might cause stress or kill the tree or provide access for pathogenic organisms. Palms that are asymptomatic for early M. h. sericeus damage could escape early visual detection and not be exposed to insecticide treatments in palm field nurseries. In such cases, a chemically-mediated lethal trap for M. h. sericeus would be extremely useful for monitoring, mass-trapping, or pathogen delivery.

ACKNOWLEDGMENTS

We thank M. Stanaland, Z. Alegria, B. Center, J. Cangiamila, and F. G. Bilz for their technical assistance, J. Perrier for preparation of Figure 1, and R. H. Schef- frahn and F. W. Howard for critically reviewing the manuscript. We are grateful to E. Suarez and P. Naranjo of Homestead, FL, and J. R. and B. Hatton of Pahokee, FL for allowing us to do ﬁeld attractancy tests on their respective properties. We thank B. Coy, D. Clinton, D. Chalot, and M. C. Thomas of the Florida Department of Agricul- ture and Consumer Services, Division of Plant Industry, D. Hull from Dade County Extension, W. L. Schall from Broward County Extension, and K. Bodman from Red- lands Research Station, Queensland for their help and suggestions. The research was supported in part by the Dade and Broward County chapters and the State or- ganization of the Florida Nursery Growers Association (FNGA). This manuscript is Florida Agricultural Experiment Stations Journal Series R-03435.

REFERENCES CITED

ARANGO S., G., AND D. RIZO. 1977. Algunas consideraciones sobre el comportamiento de Rhynchophorus palmarum y Metamasius hemipterus en caña de azucar. Revista Colombiana de Entomologia 3: 23-28. GIBLIN-DAVIS, R. M., T. J. WEISSLING, A. C. OEHLSCHLAGER, AND L. M. GONZALEZ. 1994. Field response of Rhynchophorus cruentatus (Coleoptera: Curculion- idae) to its aggregation pheromone and fermenting plant volatiles. Florida En- tomol. 77: 164-177. Giblin-Davis et al.: Trapping of Metamasius hemipterus 255

HALFPAPP, K. H., AND R. I. STOREY. 1991. Cane weevil borer, Rhabdoscelus obscurus (Coleoptera: Curculionidae), a pest of palms in Northern Queensland, Austra- lia. Principes 35:199-207. MITCHELL, G. A. 1978. The estimation of banana borer population and resistance levels. Windward Island Banana Research & Development Bulletin No. 2. Cas- tris, St. Lucia, West Indies. OEHLSCHLAGER, A. C., H. D. PIERCE, JR., B. MORGAN, P. D. C. WIMALARATNE, K. N. SLESSOR, G. G. S. KING, G. GRIES, R. GRIES, J. H. BORDEN, L. F. JIRON, C. M. CHINCHILLA, AND R. G. MEXAN. 1992. Chirality and field activity of Rhyn- chophorol, the aggregation pheromone of the American palm weevil. Natur- wissenschaften 79: 134-135. PEREZ, A. L., L. M. GONZALEZ, H. D. PIERCE, JR., A. C. OEHLSCHLAGER, G. GRIES, R. GRIES, R. M. GIBLIN-DAVIS, J. E. PEÑA, R. E. DUNCAN, AND C. CHINCHILLA. 1994. Aggregation pheromones of the sugarcane weevil, Metamasius hemipterus sericeus (Olivier). Naturwissenschaften 81: (in press). RAIGOSA, J. 1974. Nuevos diseños de trampas para control de plagas en caña de azucar (Saccharum officinarum L.). Memorias II Congreso de la Sociedad de Entomologia Colombiana, Julio 7 al 10 de 1974, Cali, Colombia. pp. 5-23. SAS INSTITUTE. 1985. SAS user's guide: statistics, 5th ed. SAS Institute, Cary, N.C. TERAN, F. O. 1968. The potential use of insecticide-treated cane pieces to attract and control adults of the sugarcane weevil, Metamasius bilobus, in Bolivia. J. Econ. Entomol. 61: 1031-1033. TILLES, D. A., G. NORDLANDER, H. NORDENHEM, H. H. EIDMANN, A. WASSGREN, AND G. BERGSTRÖM. 1986. Increased release of host volatiles from feeding scars: a major cause of field aggregation in the pine weevil Hylobius abietis (Co- leoptera: Curculionidae). Environ. Entomol. 15: 1050-1054. VAURIE, P. 1966. A revision of the Neotropical genus Metamasius (Coleoptera, Curcu- lionidae, Rhynchophorinae). Species groups I and II. Bull. American Mus. Nat. Hist. 131: 213-337. WEISSLING, T. J., R. M. GIBLIN-DAVIS, R. H. SCHEFFRAHN, AND N. M. MARBAN-MENDOZA. 1992. Trap for capturing and retaining Rhynchophorus cruentatus (Coleoptera: Curculionidae) adults using Sabal palmetto as bait. Florida Entomol. 75: 212-221. WEISSLING, T. J., R. M. GIBLIN-DAVIS, AND R. H. SCHEFFRAHN. 1993. Laboratory and field evidence for male-produced aggregation pheromone in Rhynchophorus cruentatus (F.) (Coleoptera: Curculionidae). J. Chem. Ecol. 19: 1195-1203. WEISSLING, T. J., R. M. GIBLIN-DAVIS, G. GRIES, R. GRIES, A. L. PEREZ, H. D. PIERCE, JR., AND A. C. OEHLSCHLAGER. 1994. An aggregation pheromone of the palmetto weevil, Rhynchophorus cruentatus (F.) (Coleoptera: Curculionidae). J. Chem. Ecol. 20: (in press). WOLCOTT, G. N. 1955. Entomologia economica Puertorriqueña. Univ. Puerto Rico Es- tac. Exp. Agr. Bull. 125: 1-208. WOODRUFF, R. E., AND R. M. BARANOWSKI. 1985. Metamasius hemipterus (Linnaeus) recently established in Florida (Coleoptera: Curculionidae). Florida Dept. Ag- ric. & Consumer Serv. Division of Plant Industry, Entomology Circular No. 272. 4 pp.

256 Florida Entomologist 77(2) June, 1994

ADULT SURVIVAL AND MOVEMENT IN MALES OF THE DAMSELFLY HETAERINA CRUENTATA (ODONATA: CALOPTERYGIDAE)

ALEJANDRO CÓRDOBA-AGUILAR Instituto de Ecología, A. C., Biosistemática de Insectos. Apdo. Postal 63. Xalapa, Veracruz. 91000 México

ABSTRACT

During a mark-recapture study of the territorial damselfly Hetaerina cruentata (Rambur), several changes in population size, survivorship and birth rates occurred, probably caused by physical conditions and a natural seasonal decline in population size. The average survival rate of males (expected value per day = 0.978) was the highest reported for the Calopterygidae and possibly all Odonata. This value could have been affected by factors that reduced energy expenditures at the end of the season. Analysis of male dispersion showed aggregated distributions, which are expected for territorial species. While most males dispersed, no age-class showed a tendency to move upstream or downstream. Adult females were rare; however, the sex ratio of nymphs was not significantly different from one. Key Words: Population size, survivorship, movements, age, males, damselfly.

RESUMEN

Durante un estudio de marca-captura de la libélula territorial Hetaerina cruentata (Rambur), ocurrieron varios cambios en el tamaño poblacional, sobrevivencia y tasas de nacimiento, motivados probablemente por las condiciones físicas y una dis- minución natural del lamanó publacional. La tasa de sobrevivencia masculina (valor esperado por día = 0.978) es la más alta reportada para un Calopterígido y quizás para todo el orden Odonata. Este valor pudo haber sido afectado por factores que redujeron los gastos energéticos al ﬁnal de la estación. Los análisis de la dispersión de machos indicaron una distribución agregada, lo cual está de acuerdo con el comportamiento territorial de los machos. A pesar de que los machos se dispersaron, nin- guna de las clases de edad mostró una tendencia a ir río arriba o abajo. Aunque las hembras adultas fueron raras, la proporción de sexos no fue estadísticamente signif- icativa.

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Ecological parameters frequently explain interesting biological variables. Much of community and population structure is well known today because of the study of species abundance and composition. Odonates are excellent models in population ecology, and population studies of some species have been extensive. However, most studies have been made on temperate species, and our knowledge of tropical species is still limited (Garrison & González-Soriano 1988). The American genus Hetaerina is widely distributed in tropical areas, and well studied taxonomically (see Garrison 1990). Nevertheless, diverse aspects of behavior and ecology in this genus have been poorly described (Johnson 1961, 1962, 1963, Bick & Sulzbach 1966; also see Alcock 1982 and Eberhard 1986). Considering the volume of ecological information on other species of odonates, the conspicuousness of Hetaerina and the suitability of the rivers for study, it is surprising almost no population data is available except for a few records of H. cruentata biology (e.g. William- son 1923). My aim in this study was to obtain detailed information on the population biology of a species in this interesting genus. Mark-recapture methods were used to This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.uﬂ.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

Córdoba-Aguilar: Survival and Movement in a Damselﬂy 257 study demographic parameters of a low-density population of the damselﬂy H. cruentata. Data were compared with those of other calopterygids.

MATERIALS AND METHODS

The study was carried out over 21 days (3-24 September, 1992) at the Sordo River (Fig. 1), which is situated north of Xalapa, Veracruz, Mexico (19°30'N, 96°95'W). The surrounding area is strongly disturbed by agriculture, but some areas still contain virgin forest vegetation. Other odonates found there were Brechmorhoga vivax Cal- vert (very common), Argia fissa Selys (common), Apanisagrion lais (Selys) (common), Aeshna psilus Calvert (rare), Remartinia luteipennis florida (Hagen) (rare) and a few species of Gomphidae (rare). Eighty-nine 2x2 m quadrats were made along the three branches of the river. An indelible ink pen was used to write a distinct number, never exceeding three digits, on either anterior wing. Captures and recaptures were made just once daily, except on days 7, 9, 12, 13, 22 and 23. Sampling activity started at 0900 and ended at 1200- 1230 hours. The following data were recorded for each sighting: individual number, sex, age, activity, time, quadrat and date. Individuals were placed into one of three age classes: teneral, juvenile mature and old mature. Tenerals were those individuals recently emerged. In general, they were of soft, fragile construction, difficult to manipulate and flew in a zig-zag pattern. Teneral males did not possess the same blackish color as mature adults and the red spots at the bases of the wings were not intense. In teneral females, color was not as distinct as in the other age classes, but the fragility and weak flight was easily seen. Juvenile mature (JM) animals were identified by their color (principally the red basal wing spots in males and brilliant greenish body color in females). In old mature (OM) animals, body color was well developed and the wings were opaque. Some individuals showed wing damage. A fourth type, mature (M), was frequently assigned to those individuals in transition between juvenile and old mature. Only some of the population parameters for females could be obtained because they were very scarce at the study site. Jolly's (1965) multiple capture and recapture method was used to estimate the number of marked animals in the population, population size, survivorship and the recruitment into the population. Jolly's population survivorship is a relative value ranging from 0 to 1 (values higher than 1 make no sense). The recruitment estimate is assumed to be the net increase resulting from birth and migration. Average daily survivorship (Garrison & Hafernik 1981) was also calculated by averaging daily sur-

Fig. 1. Central portion of the study site (Sordo River, Xalapa, Veracruz, Mexico). Shaded areas are rocks. Points indicate distribution of individuals.

258 Florida Entomologist 77(2) June, 1994 vivorship values. Values for missed days between recaptures were estimated as follows (Garrison & Hafernik 1981):

1/d one day survival rate = (survival rate) where, d is the number of missed days taking into account the last day of recapture. The survival rate was translated into expected life-span by the method of Cook et al. (1967), which according to the average of daily survival rate, indicates a potential expected life-span in the population Ð1 expected life-span = ------() loge average daily survival rate Velocity patterns were calculated by Scott's (1974, 1975) procedure to obtain averages of total distance travelled in meters (D), time in days (T) and velocity (V=D/T). Adult velocity is used here as the distance travelled per time by an individual. For this species, this parameter is important because changes in velocity can reﬂect differences in territorial tenure. Damselﬂy movements were separated according to the direction travelled (upstream or downstream) and the age of the individual, looking for net changes in total displacement (not daily movements). These estimates of movements and velocity were made carefully only after checking the number of days elapsed between recaptures to avoid mistakes when assigning ages. Morisita's (1959) index of dispersion, Lloyd's (1967) mean crowding index and the variance-to-mean ratio (Southwood 1979) were used to analyze population dispersion. For each method, values less than one indicate a uniform distribution, values greater than one indicate an aggregated distribution and a value of one indicates a 2 random distribution. All χ analyses include Yate's correction factor.

RESULTS

A total of 208 individuals was marked (Table 1). Recaptures differed greatly between sexes, showing, as in other studies, a greater percentage of males. An important factor in the study was that female numbers were low, probably because of a natural seasonal decline. Results referring to females must be interpreted carefully.

Population Size, Survivorship and Recruitment

The results of the estimates by Jolly's method are summarized in Table 2. There are two population peaks (see Fig. 2) on days 10 and 17 (about 1080 and 2700 individuals, Table 2). After day 17, the population decreased sharply. Population estimates probably increase on sunny days after a few days of rain, as could have been the case for days 7, 12 and 13 when strong rains fell on the river. However, this does not explain the peak on day 10 because it did not rain on day 9 (however, it should also be noted that one of the largest standard errors also occurred on day 10).

TABLE 1. RECAPTURE INDEXES OF HETAERINA CRUENTATA. N = TOTAL CAPTURED INDIVIDUALS, RC = NUMBER OF INDIVIDUALS RECAPTURED, %RC = PRO- PORTION OF MARKED INDIVIDUALS RECAPTURED. Sex N RC %RC Male 182 118 64.8 Female 26 6 23.07 Totals 208 124 59.6

Córdoba-Aguilar: Survival and Movement in a Damselﬂy 259

TABLE 2. POPULATION PARAMETERS OF HETAERINA CRUENTATA ESTIMATED PER DAY BY JOLLY METHOD (1965). M = MARKED POPULATION ESTIMATES, N = POP- ULATION SIZE, SURV = SURVIVORSHIP, B = NUMBER OF ANIMALS ENTERING INTO POPULATION, AND SE = STANDARD ERRORS. Days M N±SE Surv±SE B 3 ------4 291 385±34 1.44±0.18 64 5 437 621±101 0.95±0.15 -4 6 427 588±104 0.77±0.09 41 8 341 496±58 2.61±0.71 -216 10 933 1083±295 0.35±0.09 73 11 337 462±61 2.44±0.79 -344 14 849 786±242 0.86±0.32 219 15 732 898±192 1.54±0.64 -98 16 1145 1288±479 1.91±1.27 237 17 2204 2708±1511 0.57±0.38 46 18 1279 1611±608 0.45±0.21 162 19 582 888±280 1.25±0.69 -141 20 757 976±465 0.38±0.23 23 21 293 395±162 ------24 ------

Survivorship fluctuated greatly. On several occasions, values higher than one were obtained; such values make no biological sense. The daily survival rate reflects the differences in the number of recaptures. High recapture rates were associated with high daily survival rates. The estimated survival curve for males captured on the first day, is shown in Fig. 3. This survival curve included individuals of all ages except tenerals (JM n = 9, M n = 7, OM n = 43). The curve clearly shows a decline throughout the study. No analysis of females was made due to their infrequent recaptures. Average survival rate was 0.978 per day, which, when converted to life expectancy by Cook et al.'s method is 44.9 days. This result is compared with the survivorship and life expectancy of males of other species in Table 3.

Dispersion, Movements and Velocity

Morisita's index of dispersion for males was 3.63 which is statistically different from one (p < 0.05). Lloyd's mean crowding index was 2.19±0.01 (S.E.) while the variance-to-mean ratio (Southwood 1978) was 2.75. All three indices indicate aggregated distributions. Aggregations of individuals in the central portion of the study site can be seen in Fig. 1. The same number of JM males moved up the river as down. In M males, more in- 2 dividuals moved downstream, but the difference was not significant (χ = 0.02, d.f. = 1, p > 0.05). More OM males moved upstream than down, but this difference was also 2 not significant (χ = 0.93, d.f. = 1, p > 0.05). There was, however, a significant difference when the number of individuals of each age dispersing was compared with those remaining in the same quadrat among recaptures. More OM and JM males re- 2 mained in the same quadrats (or very close) (χ2 = 16.6, d.f. = 1, p < 0.001 for OM, χ = 5.66, d.f. = 1, 0.05 > p > 0.01 for JM). Nevertheless, among M individuals none remained on the same quadrat for all recaptures (n = 25). Although there were more animals (regardless of age) that moved upstream (64%) than downstream (36%), the 260 Florida Entomologist 77(2) June, 1994

Fig. 2. Estimated population size in Hetaerina cruentata as estimated by Jolly (1965) method in the study. Vertical lines are standard errors.

TABLE 3. AVERAGE SURVIVORSHIP COMPARISONS AMONG MALES IN SEVERAL SPECIES OF CALOPTERYGIDAE AND THEIR EXPECTED VALUE IN DAYS. 1 = CORDERO (1989), 2 = CONRAD & HERMAN (1990), 3 = HIGASHI (1976 IN CORDERO, 1989), 4 = NOMAKUCHI ET AL. (1988), AND 5 = THIS STUDY. ESTIMATES IN PARENTHESES IN CALOPTERYX HAEMORROIDALIS AND MNAIS PRUINOSA (4) REFER TWO COMPARATIVE AVERAGES. Species Avg Days Calopteryx virgo (1) 0.86 6.6 C. haemorroidalis (1) 0.94 (0.91) 16.2 (10.6) C. xanthostoma (1) 0.66 2.4 C. aequabilis (2) 0.767 3.8 Mnais pruinosa (3) 0.943 ------M. pruinosa (4) 0.944 (0.947) 17.6 (18.4) Hetaerina cruentata (5) 0.978 44.9 Córdoba-Aguilar: Survival and Movement in a Damselﬂy 261

Fig. 3. Estimated percent of survivorship in males of Hetaerina cruentata captured in the first day of study (n = 59). difference was not significant (χ2 = 1.05, d.f. = 1, p > 0.05). Movement patterns of tenerals and females could not be estimated. Scott's velocity indices were as follows: time in days between first and last recapture in males was 11.4, mean of total distance travelled in m was 44.3, and the mean velocity was 3.9 m per day. For individual velocities, males only and different age classes were analyzed separately. The procedure used was to take the quadrat of the first recapture (after capture) and calculate the number of days between captures. This short interval among captures meant age did not change much between captures. There were no velocity differences among age classes (one-way ANOVA, p > 0.05). Distances travelled between males and females differed significantly (t test = 2.13, d.f. = 29, p < 0.025). Females had higher velocity patterns as in other species of Calopteryx (see Cordero 1989, Conrad & Herman 1990, Waage 1972). The differences among tenerals and the other age classes is probably great because a teneral was never recaptured within 15 m of the river. Two males travelled 66.9 m between suc- cessive recaptures while other males remained in the same quadrats for several days. This fact has been reported previously for other calopterygid species [He- 262 Florida Entomologist 77(2) June, 1994 taerina americana (Fabricius), Johnson 1962; H. vulnerata, Alcock 1982; H. macropus Selys, Eberhard 1986].

Sex Ratio and Mortality

Operational sex ratio always was notably skewed toward males (182 males, 26 females), but these estimates do not reflect the primary sex ratio and result mainly from behavioral differences between the sexes. Estimates of sex ratio were obtained also by sexing larvae. In spite of a slight deviation toward females (43 females, 36 males), the ratio was not significantly different from 1:1 (χ2 = 0.455, d.f. = 1, p > 0.05). During the study, no attacks by vertebrates like birds, frogs or fishes were noted although they are usually regarded as common predators of other odonates. On one occasion an ovipositing female was attacked by a giant aquatic bug (Family Belosto- matidae). Some newly emerged individuals were found in spider webs. Although the sampling quadrats increased spider density by providing new, favorable places for webs, no adults of H. cruentata were observed in these webs.

DISCUSSION

Population size (Fig. 2) may have changed during the study as a result of strong rains, which affected afternoons and occasionally whole days. The rains may also have been responsible for the highly fluctuating survivorship (Table 2). However, other reasons exist for these changes (predation, migration, among others) that could not be measured. As the study progressed, captures decreased and this affected several additional parameters, including the Jolly's estimates (especially the S.E.'s). Cordero (1989) found that recaptures (% RC) in three species of Calopteryx (haemorroidalis, xanthostoma and virgo L.) did not exceed 53% for both sexes. The highest percentage was for C. virgo (54%). In this respect, H. cruentata males showed a relatively high percentage (64.8%, Table 1) of recaptures. The highest frequency of recaptures for a damselfly is that of Ischnura gemina (Kennedy) (Garrison & Hafernik 1981) with 84% for both sexes and 90% for males alone. In addition, if it is assumed that during the study the population was at low density when compared with other months (pers. obs.), then the high recapture rates (higher than other species of Calopterygidae) could result because males are better able to successfully de- fend territories for several days when there is relatively little competition for sites. Comparisons within the calopterygids and with other species are needed to test this hypothesis. Primary sex ratio was not different from one in H.cruentata. The same occurs in other members of the calopterygids as Calopteryx virgo, C. haemorroidalis, C. xanthostoma, C. virgo (Cordero 1989), C. maculata (Beauvois), C. dimidiata (Waage 1980). Waage (1980) has pointed out that the sex ratio should be 1:1 among all odonates. Survivorship estimated by Jolly's method was quite variable (Table 2), but shows a general decline over the month. Some of the variation probably results from including several age classes in the analysis. The curve declines mainly due to the mortality of OM animals. JM individuals presumably experience different mortality pressures. It is interesting to note that the survival rate by Cook et al.'s method was the highest (44.9 days) in the suborder, and perhaps for odonates in general. Corbet (1958) has pointed out that adult life-span of neotropical damselflies is longer compared with those of temperate latitudes, presumably due to less variable abiotic conditions in the tropics. However, reduced energetic expenditures may have also lengthened life in H. cruentata. These factors could include reduced activity in rainy Córdoba-Aguilar: Survival and Movement in a Damselfly 263 days because of cooler temperatures or reduced competition for territories due to low density conditions. H. cruentata is a territorial species with behavioral biology similar to other calopterygids (Córdoba-Aguilar 1994). Several males perch, aggressively defending certain areas (oviposition sites) against other males, where females arrive, searching for copulation and oviposition. These sites have high male density in relation to others where oviposition sites do not exist. Under low density conditions, movement of mature males are minimal, and presumably male velocity diminishes due to the availability of many defensible territories, as has been demonstrated in Calopteryx aequabilis Say at low density by Conrad & Herman (1990). Likewise, when density is modified, other behavioral parameters are modified too. For example, fights among territorial males in Mnais pruinosa Selys at the transitional period (when there were mature and immature individuals in the population and at, presumably, low density) influenced movements on the stream of individuals that could not win a territory (Nomakuchi & Higashi 1985). Once these males established their territories, and density was higher, they moved infrequently (Nomakuchi & Higashi 1985). This result was similar to territorial males of H. cruentata. However, other conditions can influence dispersal and movements. In general, animals experiencing poor feeding or mating opportunities are usually more likely to disperse than those with good conditions (Lomnicki 1978, Lawrence 1987). On the other hand, the few differences among age classes in movements and velocities are interesting. Eberhard (1986) observed that Hetaerina macropus Selys pe- riodically left the river but had a low tendency to disperse, in general. Cordero (1989) found that in Calopteryx haemorroidalis, juvenile males moved greater distances than mature males, and movements in C. xanthostoma were up river probably resulting from strong winds. Likewise, Higashi & Ueda (1982) found similar results in a greater percentage of individuals of C. cornelia (Selys). Nevertheless, H. cruentata males did not display significant upstream or downstream movements. More studies are needed to examine dispersal at high densities.

ACKNOWLEDGMENTS

Special thanks to F. Capistrán-Hdez., E. Estrada-C. and S. Bedoy-G. for helping in the field work. Financial support was obtained as a part of a related project in Ischnura denticollis from Sigma Xi, The Scientific Research Society. Thanks to I. Chamorro-F. for her constant support in the field data analysis, to R. Novelo-Gtz. (IEX) and E. González-S. (IBUNAM), for their criticisms and help with translation to English. Finally, my deep appreciation to K. F. Conrad (Canada) and S. D. Porter (U.S.A.) for extensive suggestions that greatly improved the manuscript.

REFERENCES CITED

ALCOCK, J. 1982. Post-copulatory mate guarding by males of the damselfly Hetaerina vulnerata (Odonata: Calopterygidae). Anim. Behav. 30: 99-107. BICK, G. H., AND D. SULZBACH. 1966. Reproductive behavior of the damselfly, He- taerina americana (Fabricius) (Odonata: Calopterygidae). Anim. Behav. 14: 156-158. CONRAD, K. F., AND T. B. HERMAN. 1990. Seasonal dynamics, movements and the effect of experimentally increased female densities on a population of imaginal Calopteryx aequabilis (Odonata: Calopterygidae). Ecol. Entomol. 15: 119-129. COOK, L. M., P. P. BROWER, AND H. J. CROZE. 1967. The accuracy of a population estimation from multiple recapture data. J. Anim. Ecol. 36: 57-60. CORBET, P. S. 1958. Temperature in relation to seasonal development of British dragonflies (Odonata). Proc. Tenth Int. Congr. Entomol. 2: 755-758. 264 Florida Entomologist 77(2) June, 1994

CORDERO, A. 1989. Estructura de tres comunidades de Calopteryx (Odonata: Calop- terygidae) con diferente composición específica. Limnética 5: 83-91. CÓRDOBA-AGUILAR, A. 1994. Infuencia del tomaño coporal y la residencialidad sobre el éxito en las contiendas maculinas en la libélula Hetaerina Cruentata (Ram- bur) (Odonata: Calopterygidae). Bachelor thesis. Universidad Veracruzana. Mexico. EBERHARD, W. G. 1986. Behavioral ecology of the tropical damselfly Hetaerina macropus Selys (Zygoptera: Calopterygidae). Odonatologica 15: 51-60. GARRISON, R. W. 1990. A synopsis of the genus Hetaerina with descriptions of four new species (Odonata: Calopterygidae). Trans. Am. Ent. Soc. 116: 175-259. GARRISON, R. W., AND E. GONZALEZ-SORIANO. 1988. Population dynamics of two sib- ling species of neotropical damselflies, Palaemnema desiderata Selys and P. paulitoyaca Calvert (Odonata: Platystictidae). Folia Entomol. Mexicana 76: 5- 24. GARRISON, R. W., AND J. HAFERNIK, JR. 1981. Population structure of the rare damselfly, Ischnura gemina (Kennedy) (Odonata: Coenagrionidae). Oecologia 48: 377-384. HIGASHI, K., AND T. UEDA. 1982. Territoriality and movement pattern in a population of Calopteryx cornelia (Selys) (Zygoptera: Calopterygidae). Odonatologica 11: 129-137. JOHNSON, C. 1961. Breeding behavior and oviposition in Hetaerina americana Fabr- icius and H. titia (Drury) (Odonata: Agriidae). Canadian Entomol. 93: 260- 266. JOHNSON, C. 1962. A description of the territorial behavior and a quantitative study of its function in males of Hetaerina americana (Fabricius) (Odonata: Agri- idae). Canadian Entomol. 94: 178-190. JOHNSON, C. 1963. Interspecific territoriality in Hetaerina americana (Fabricius) and H. titia (Drury) (Odonata:Calopterygidae) with a preliminary analysis of the wing color pattern variation. Canadian Entomol. 95: 575-582. JOLLY, G. M. 1965. Explicit estimates from capture-recapture data with both death and immigration-stochastic model. Biometrika 52: 225-247. LAWRENCE, W. S. 1987. Dispersal: an alternative mating tactic conditional on sex ratio and body size. Behav. Ecol. Sociobiol. 21: 367-373. LLOYD, M. 1967. “Mean crowding”. J. Anim. Ecol. 36: 1-30. LOMNICKI, A. 1978. Individual differences between animals and the natural regula- tion of their numbers. J. Anim. Ecol. 47: 461-465. MORISITA, M. 1959. Measuring the dispersion of individual and analysis of the distri- butional patterns. Mem. Fac. Sci. Kyushu Univ. 2: 215-235. NOMAKUCHI, S., AND K. HIGASHI. 1985. Patterns of distribution and territoriality in the two male forms of Mnais pruinosa Selys (Zygoptera: Calopterygidae). Odo- natologica 14: 301-311. NOMAKUCHI, S., K. HIGASHI, AND M. MAEDA. 1988. Synchronization of reproductive period among two male forms and female of the damselfly Mnais pruinosa Selys (Zygoptera: Calopterygidae). Ecol. Res. 3: 75-87. SCOTT, J. A. 1974. Adult behavior and population biology of two skippers (Hesperi- idae) mating in contrasting topographic sites. J. Res. Lepidoptera 12: 181-196. SCOTT, J. A. 1975. Flight patterns among eleven species of diurnal Lepidoptera. Ecol- ogy 56: 1367-1377. SOUTHWOOD, T. R. E. 1979. Ecological methods. Chapman & Hall. London. WAAGE, J. K. 1972. Longevity and mobility of adult Calopteryx maculata (Beauvois, 1805) (Zygoptera: Calopterygidae). Odonatologica 1: 155-162. WAAGE, J. K. 1980. Adult sex ratio and female reproductive potential in Calopteryx (Zygoptera: Calopterygidae). Odonatologica 9: 217-230. WILLIAMSON, E. B. 1923. Notes on the habitats of some tropical species of Hetaerina. Publ. Michigan St. Univ. Mus. 130: 1-46.

Childers: Brevipalpus Feeding Injury & Control on Citrus 265

FEEDING INJURY TO 'ROBINSON' TANGERINE LEAVES BY BREVIPALPUS MITES (ACARI: TENUIPALPIDAE) IN FLORIDA AND EVALUATION OF CHEMICAL CONTROL ON CITRUS

CARL C. CHILDERS Citrus Research and Education Center University of Florida, IFAS 700 Experiment Station Road Lake Alfred, FL 33850

ABSTRACT

Larvae, nymphs and adults of the false spider mites Brevipalpus phoenicis (Gei- jskes) and B. obovatus Donnadieu (Acari: Tenuipalpidae) were found feeding on the ventral surface along the midrib of 'Robinson' tangerine leaves during December 1990. Resinous irregular areas were present in association with the chlorotic midvein areas of injured cells on the lower leaf surface. Similar injury was occasionally evident on some lateral veins in association with the mites. Yellowed, blistered areas were evident on the upper leaf surface opposite the injured areas on the lower leaf. Leaf drop was evident only on trees that were heavily infested with false spider mites. This type of leaf injury by Brevipalpus mites has not been previously reported on Florida citrus. Chemical control evaluations showed that AC 303,630 in combination with petroleum oil, pyridaben, fenbutatin-oxide, dicofol or high rates of sulfur provided at least 35 days control. Ethion was less effective and carbaryl failed to control these mites. Key Words: Tenuipalpidae, injury, false-spider mites.

RESUMEN

En diciembre 1990, se encontraron larvas, ninfas y adultos del acaro falso Brevi- palpus phoenicis (Geijskes) y B. ovatus Donnadieu (Acari: Tenuipalpidae) alimentan- dose en el envéz de las hojas de mandarina "Robinson". Se observaron areas resinosas irregulares conjuntamente con areas cloroticas en la nervadura central en el envéz. Un daño similar se encontró ocasionalmente en algunas nervaduras se- cundarias. En el haz se observó, amarillamiento y ampollamiento de las areas afecta- das. Se observó defoliación unicamente en aquellos arboles con una gran infestación de acaros falsos. Este tipo de daño no ha sido reportado en cítricos en Florida. Las evaluaciones de control químico demostraron que AC 303,630 en combinación con aceite, pyridaben, fenbutatin-oxide, dicofol o altas dosis de azufre brindaban control por 35 días. Ethion fué menos efectivo y carbaryl no controló estos acaros.

————————————

Three species of false spider mites [Brevipalpus californicus (Banks), B. obovatus Donnadieu, and B. phoenicis (Geijskes) (Acari: Tenuipalpidae)] have been collected from citrus in Florida (Denmark 1984). All three species are cosmopolitan and occur on citrus in Asia, Africa, Australia, Europe, the Middle East, South America, and the United States (Jeppson et al. 1975). These mites feed on fruit, stems, branches, and leaves of citrus. On leaves, they are most commonly found on the lower surface near the midrib or veins. False spider mites are reddish in color, slow moving, and not readily detected because of their small size and sluggish behavior. They are about 260 µm in length (Muma 1961). Brevipalpus mites are found on many perennial plants and have relatively long life cycles compared with other phytophagous mite pests, especially on

266 Florida Entomologist 77(2) June, 1994 citrus. Population levels of these mites tend to increase slowly due to their relatively long life cycle (i.e., 35 days at temperatures between 70 and 85°F) (Manglitz & Cory 1953). The greatest concern with Brevipalpus species on citrus in Florida and elsewhere in the world has been their association with the viral disease, leprosis (Chiavegato et al. 1982). Leprosis and infestations of B. californicus very nearly destroyed the citrus industry in Florida before the late 1920's (Knorr et al. 1968). Leprosis was first observed in Florida in the 1860's and ultimately was found in 17 counties that essen- tially represented the total citrus growing area within Florida at that time. The problem disappeared from Florida citrus in the late 1920's apparently following the widespread usage of sulfur for mite control (Knorr et al. 1968). Symptoms of leprosis have been reported to occur on fruit, leaves, shoots and large limbs. Chestnut-brown spots from pinhead size to 6 mm in diameter occur on oranges (Knorr 1973). In Florida, this malady has been referred to as nailhead rust (Knorr & Price 1958). Similar lesions occur on both upper and lower leaf surfaces, especially along the margins. Scaly lesions, called scaly bark, grow on the twigs (Knorr & Price 1958). Chiavegato et al. (1982) conducted transmission studies with the virus causing leprosis and its vector B. phoenicis. The causal agent of leprosis is presumed to be a mite-vectored bacilliform virus (Garnsey et al. 1988). Leprosis has been transmitted by larvae of B. phoenicis after a 24 h acquisition period but nymphs and adults were less efficient (Chiavegato & Salibe 1984). According to Knorr (1959), only B. californicus has been associated with this disease in Florida. B. obovatus has been associated with leprosis in Argentina and Ven- ezuela (Garnsey et al. 1988). B. phoenicis is the vector in Brazil (Chiavegato et al. 1982). Knorr et al. (1968) found foliar and fruit lesions on 'Valencia' orange that were similar in color, size and diameter to those caused by leprosis. However, corresponding lesions on shoots, twigs or branches were not evident. Large populations of B. californicus and B. phoenicis were associated with a rind spotting of grapefruit in Texas between June and October 1966. The injury was only on fruit and resembled leprosis-like spotting (Dean & Maxwell 1967). Brevipalpus gall or nodal galling resulted from B. phoenicis feeding on citrus seedlings in Venezuela and Florida (Knorr & Denmark 1970). Plants subsequently died since they were unable to leaf out. They further reported leaf drop on sour orange seedlings where B. phoenicis occurred in association with the fungus Elsinoe fawcetti Bitancourt & Jenkins. Another foliar problem called phoenicis blotch has been reported in association with B. phoenicis on Florida citrus. Diffuse chlorotic spotting of foliage in sweet orange trees occurred which resembled early stages of leprosis. However, no gumming of the affected areas occurred (Knorr et al. 1960). Because little attention has been given to Brevipalpus mites in recent years, it is appropriate to review chemical control of these potentially serious pests. These mites have been observed in many citrus groves throughout Florida in conjunction with other research activities (unpub. data). A serious leaf drop problem caused by high populations of B. phoenicis and B. obovatus was identified in 'Robinson' tangerine. In- jury to leaves and subsequent chemical control evaluations are reported in this paper.

MATERIALS AND METHODS

A 'Robinson' tangerine grove located in Haines City, Polk County, Florida was vis- ited on 28 December 1990. The grower had complained about extensive leaf drop to his 3-year-old trees. Several trees were examined. Only Brevipalpus mites were found associated with leaf injury. They were removed for slide mounting in Hoyer's

Childers: Brevipalpus Feeding Injury & Control on Citrus 267 medium (Krantz 1978) and subsequent identification to species. No other arthropods were found on the leaves. Chemical control of Brevipalpus mites was assessed during 1991 and 1992. Both experiments were established in a 'Hamlin' orange grove of 20+-year-old trees in Hardee County that were 2.4 to 3.7 m high with 3 to 4.6 m canopy diameters and infested with moderate numbers of Brevipalpus mites. Tree spacing was 3.7 by 8.5 m with 321 trees per hectare. Pesticide treatments were applied dilute in 5,574 liters of water per ha with a handgun using a truck-mounted sprayer at 350 psi on 3 July 1991. The 1992 field experiment was established in the same grove site. Treatments were applied dilute in 7,781 liters of water per ha with a handgun on 1 July 1992. Experimental miticides included AC 303,630 [4-bromo-2-(4-chlorophenyl)-1- (ethoxymethyl)-5-(trifluoromethyl) pyrrole-3-carbonitrile] (American Cyanamid Corp., Princeton, NJ) formulated as a 24% emulsifiable concentrate (EC) and a 22.5 g per liter (3 lb per gallon) soluble concentrate (SC), pyridaben formulated as 20 and 75% wettable powders (WP) (BASF Corp., Research Triangle Park, NC), and Mi- crothiol sulfur (ELF ATOCHEM Corp., Tifton, GA) was a micronized WP formulation containing 80% sulfur while the other sulfur compound was a 90% WP formulation. The remaining pesticides included FC435-66 petroleum oil (Orchex  796), fenbutatin oxide (= Vendex ), dicofol 4EC, ethion 4EC, and carbaryl 80S that are registered for use on citrus. Treatments were assigned at random to single tree plots in 1991 and in a randomized complete block design in 1992 and replicated 5 times in both experiments. Each sample tree was a vigorous, healthy tree representative of the block, and each plot was separated from adjacent plots by at least one tree within and between rows. Forty fruit were picked at random around the perimeter of each sample tree after 42 d post-treatment on 14 August 1991. Each fruit was agitated in a bucket containing 80% ethyl alcohol, detergent, and bleach that was modified by increasing alcohol concentration in a mixture developed by Gilstrap & Browning (1983). The solution from each sample tree was poured into a separate labeled jar, returned to the laboratory, and then poured into a gridded Petri dish and examined using a stereomicro- scope. The number of motile Brevipalpus mites per sample was counted. In 1992, 20 fruit per sample tree were washed individually in a bucket containing the alcohol-detergent-bleach solution and vigorously shaken immediately after pick- ing. The solution from each sample tree was poured into a separate labeled jar and returned to the laboratory where the contents were counted as before. Preliminary sampling indicated that this method captured at least 95% of false spider mites on individual fruit. A subsample of 10 to 20 mites from each of 2 or 3 water-sprayed trees was slide-mounted and identified to species from sample dates during both years. The mite count data in each treatment were subjected to Log10 (X + 1) transfor- mations for statistical analysis using PROC ANOVA (SAS Institute 1991). Untrans- formed counts are presented for comparison in the tables. If the difference between treatments was significant (P ≤ 0.05), Duncan's (1955) new multiple range test was used to separate treatment means.

RESULTS AND DISCUSSION

Brevipalpus phoenicis and B. obovatus were identiﬁed from larvae, nymphs and adults collected from the injured 'Robinson' tangerine leaves in Haines City (Fig. 1A and B). Yellow blistered areas on the upper leaf surface occurred along the lengths of most infested leaves. Resinous irregular injured areas were present in association with the chlorotic midvein areas of injured cells on the lower leaf surfaces. No indication of mesophyll collapse (Pratt 1958) was evident. Similar injury was occasion-

268 Florida Entomologist 77(2) June, 1994

Fig. 1. (A) Brevipalpus sp. nymph. (B) Brevipalpus sp. adult. (C) Feeding injury caused by B. phoenicis and B. obovatus. (D) Lower leaf surface feeding injury primarily along the midvein with some lateral vein injured areas (IA) and yellow blistered area (BA) on the upper leaf surface. ally evident on some lateral veins too. Considerable leaf drop of mite infested leaves was evident (Fig. 1C and D) while adjacent non-infested 'Robinson' trees did not have injured leaves or leaf drop. All of the mites were found on the ventral surface of each infested leaf, mostly along the midvein, with motile numbers often exceeding 50 per leaf. Occasionally, chlorotic or resinous areas between lateral veins on the lower leaf surface were found infested with these mites. Mite infestations were heavier along the four or five rows on the north edge of the grove during December 1990. Leaf drop was confined to these trees. This type of leaf injury by Brevipalpus mites has not been previously reported on Florida citrus. None of the types of injury previously described from other countries corresponded to foliar injury and leaf drop reported here. Three adult B. phoenicis and 5 adult B. obovatus specimens were identified from a series of 20 prepared slides from the 'Robinson' tangerine trees in 1990. In the 1991 miticide experiment on 'Hamlin' orange, B. phoenicis and B. obovatus accounted for 68 and 32%, respectively, of the 34 specimen identifications. Slide-mounted specimens identified from subsamples collected between 9 July and 27 August 1992 were 86% (N = 139) B. phoenicis and 14% (N = 23) B. obovatus. According to Denmark (1984), B. obovatus has rarely been found on Florida citrus. B. phoenicis was the principal false spider mite observed during this study on both 'Robinson' tangerine and 'Hamlin' orange in Central Florida. B. californicus was not recorded during this study. In 1991, AC 303,630 at both rates, pyridaben 75 WP and dicofol all provided significantly better control compared with the water sprayed trees and Microthiol sulfur after 42 days (Table 1). Childers: Brevipalpus Feeding Injury & Control on Citrus 269

TABLE 1. NUMBERS OF BREVIPALPUS PHOENICIS (GEIJSKES) AND B. OBOVATUS DON- NADIEU ON ‘HAMLIN’ ORANGE FRUIT TREATED ON 3 JULY 1991. Post-treatment Means of Motile Mites per 40 Fruit1 Rate per 378 14 Aug Treatment and Formulation Liters +42 d AC 303,630 24% EC 100 ppm 3.6 c AC 303,630 24% EC 200 ppm 2.6 c Pyridaben 20 WP 181 g 8.2 bc Pyridaben 75 WP 48 g 4.8 c Dicofol 4 EC 355 ml 1.0 c Microthiol sulfur 80% WP 1.8 kg 21.8 ab Water spray — — 34.4 a

F = 6.12, df = 6,28, P = 0.0003

1Means within a column followed by the same letter are not signiﬁcantly different by ANOVA followed by Duncan’s NMRT.

In 1992, both rates of AC 303,630 + petroleum oil, fenbutatin-oxide, and both rates of sulfur provided significantly better control of the mites through 35 days post- treatment compared with the other treatments (Table 2). Pyridaben and ethion provided significantly better control of Brevipalpus spp. compared with the water- sprayed check trees; carbaryl was ineffective. The new miticides (i.e., AC 303,630 and pyridaben) were compared against established standards (i.e., sulfur, fenbutatin-oxide, dicofol) for control of Brevipalpus mites on Florida citrus. B. phoenicis has been reported to be susceptible to sulfur, dicofol and chloroben- zilate while organophosphate and carbamate pesticides are ineffective (Jeppson et al. 1975). Results from these studies confirm that carbaryl was ineffective in controlling Brevipalpus mites. Ethion showed activity against these mites; however, it was not as effective as AC 303,630, sulfur or fenbutatin-oxide. Miticide evaluations for control of B. phoenicis on tea in Kenya showed that flucythrinate, dicofol, omethoate, dimethoate, and permethrin were also effective. However, significant yield increases were obtained only in the flucythrinate and permethrin treatments (Sudoi 1990). In Brazil, dicofol at 37 g AI per 6 liters of water, clofentezine at 9.45 g AI per 6 liters of water and RU-1000 (an experimental pyrethroid) at 1.76 or 2 g AI per 6 liters of water were effective at least 35 days following treatment in controlling B. phoenicis on citrus (Mariconi et al. 1989). Cyhexatin at 20 g AI per 100 liters, binapacryl at 50 g AI per 100 liters and bifenthrin at 5.6 g AI per 100 liters were most effective in controlling B. phoenicis on citrus in Brazil (Arashiro et al. 1988).

ENDNOTE

Thanks to Gregory Evans for veriﬁcation of mite species. Technical assistance provided by Paul M. Keen, Jr., Michael G. Warmington, and Deanna K. Threlkeld was greatly appreciated. Florida Agricultural Experiment Station Journal Series No. R-03583. 270 Florida Entomologist 77(2) June, 1994 7.2 a 4.2 ab 1.0 bc 0.8 bc 3.6 ab 4.1 bc 4.4 abc ± ± ± ± ± ± ± +57 d 27 Aug = 17.93, = 0.0035 df = 8,36 1 F P 8.6 a 8.2 8.2 a 26.8 0.6 c 1.8 0.8 b 8.6 7.4 b 4.6 0.6 c 7.2 0.9 c 0 c 0.9 c 0 c ± ± ± ± ± ± ± ± +35 d 5 Aug = 11.50, = 0.0001 df = 8,36 F P

5.6 a 40.8 12.1 a 49.8 0.6 c 0 c 1.4 2.0 b 4.6 0.8 c 11.4 ± ± ± ± ± +23 d 24 Jul = 15.71, = 0.0001 ORANGE df = 8,36 SE of Motile Mites per 20 Fruit F ’ P ± AMLIN ‘H 5.4 a 44.4 0.4 b 0 c 1.2 0.8 b 4.2 0.2 b 0.8 1.7 a 48.8 ON

± ± ± ± ± +15 d 16 Jul = 22.15, = 0.0001 df = 8,36 F P ONNADIEU Post-treatment Means D 2.5 a 17.0 0.4 cd 0 b 0.6 0.9 cd 0.4 0.9 bcd 0.8 1.8 bc 0.2 0.4 ab 9.8 0.8 cd 0 b 0 c 1.4 0.4 cd 0 b 0 c 1.4 +7 d = 5.81, 8 Jul ± ± ± ± ± ± ± ± = 0.0001 df = 8,36 F P OBOVATUS . B

1 AND ) 11 a 11 0.6 10 a 9.6 24 a 1.4 29 a 2.0 25 a 3.0 12 a 0 d 0 b 0 c 1.2 13 a 4.0 14 a 0.8 12 a 0.8 ± ± ± ± ± ± ± ± ± -15 d = 0.05, 15 Jun Means = 0.9999 df = 8,36 F P EIJSKES Pre-treatment (G

Hectare PHOENICIS Rate per

1992. JULY 1 ON

REVIPALPUS B

TREATED

UMBERS FRUIT Means within a column followed by the same letter are not signiﬁcantly different by ANOVA followed by Duncan’s NMRT. followed by Duncan’s ANOVA Means within a column followed by the same letter are not signiﬁcantly different 1 Water spray Water (check) — — 50 Sulfur 90 WP 56 kg 45 Sulfur 90 WP 28 kg 59 EthionEC 4 liters7 66 Pyridaben 75 WP g 119 62 Fenbutatin-oxide 50 WP 2.2 kg 49 Carbaryl 80 S 5.6 kg 49 AC 303,630+ Orchex 796 oil FC 435-66 3 SC 0.5% 200 ppm 51 Treatment and Formulation Treatment AC 303,630 + Orchex 796 oil FC 435-66 3 SC 0.5% 100 ppm 48 TABLE 2. N Childers: Brevipalpus Feeding Injury & Control on Citrus 271

REFERENCES CITED

ARASHIRO, F. Y., A. J. RAIZER, C. A. SUGAHARA, R. MOTA, J. M. SILVA, AND F. A. M. MARICONI. 1988. New ﬁeld tests for chemical control of the citrus leprosis mite Brevipalpus phoenicis (Geijskes, 1939) on orange trees. Anais da Escola Sup. Agric. Luiz de Queiroz 45: 67-78. CHIAVEGATO, L. G., M. M. MISCHAN, AND M. A. SILVA. 1982. Prejuizos e transmissi- bilidade de sintomas de leprose pelo acaro Brevipalpus phoenicis (Geijskes, 1939) Sayed, 1946 (Acari, Tenuipalpidae) em citros. Cientiﬁca 10: 265-271. CHIAVEGATO, L. G., AND A. A. SALIBE. 1984. Transmissibility of leprosis symptoms by Brevipalpus phoenicis to young citrus plants under laboratory conditions, pp. 218-221 in S. M. Garnsey, L. W. Timmer, and J. A. Dodds [eds.] Compendium of citrus diseases. Proc. Conf. Int. Organ. Citrus Virol. 9th. IOCV, Riverside, California. DEAN, H. A., AND N. P. MAXWELL. 1967. Spotting of grapefruit as associated with false spider mites. Proc. Rio Grande Valley Hort. Soc. 21: 35-45. DENMARK, H. A. 1984. Brevipalpus mites found on Florida citrus (Acarina: Tenuipal- pidae). Florida Dept. Agric. & Consumer Serv., Div. Plant Ind. Entomol. Circ. 69. DUNCAN, D. B. 1955. Multiple range and multiple F test. Biometrics 11: 1-42. GARNSEY, S. M., C. M. CHAGAS, AND L. G. CHIAVEGATO. 1988. Leprosis and zonate chlorosis, pp. 43-44 in J. O. Whiteside, S. M. Garnsey, and L. W. Timmer [eds.] Compendium of citrus diseases. APS Press, The American Phytopathological Society, St. Paul, Minn. GILSTRAP, F. E., AND H. W. BROWNING. 1983. Sampling predaceous mites associated with citrus. Texas Agric. Exp. Sta. PR-4139. 5 p. JEPPSON, L. R., H. H. KEIFER, AND E. W. BAKER. 1975. Mites injurious to economic plants. Univ. California Press, Berkeley. 614 p. KNORR, L. C. 1959. Presenting Brevipalpus mites and some questions that they pose. Citrus Mag. 21: 8-12, 20, 22. KNORR, L. C. 1973. Citrus diseases and disorders, an alphabetized compendium with particular reference to Florida. Univ. Presses of Florida, Gainesville. 163 p. KNORR, L. C., AND H. A. DENMARK. 1970. Injury to citrus by the mite Brevipalpus phoenicis. J. Econ. Entomol. 63: 1996-1998. KNORR, L. C., H. A. DENMARK, AND H. C. BURNETT. 1968. Occurrence of Brevipalpus mites, leprosis, and false leprosis on citrus in Florida. Florida Entomol. 51: 11- 17. KNORR, L. C., AND W. C. PRICE. 1958. Leprosis, pp. 112-114 in R. M. Pratt [ed.], Flor- ida guide to citrus insects, diseases and nutritional disorders in color. Agric. Exp. Sta., Gainesville, FL. KNORR, L. C., B. N. WEBSTER, AND G. MALAGUTI. 1960. Injuries to citrus attributed to Brevipalpus mites, including Brevipalpus gall, a newly reported disorder in sour-orange seedlings. FAO Plant Prot. Bull. 8: 141-148. KRANTZ, G. W. 1978. A manual of acarology. 2nd ed. Oregon State Univ. Book Stores, Inc., Corvallis. MANGLITZ, G. R., AND E. N. CORY. 1953. Biology of Brevipalpus australis. J. Econ. Entomol. 46: 116-119. MARICONI, F. A. M., R. C. RANGEL, R. HAMAMURA, A. I. CLART, L. F. MESQUITA, M. A. C. CARDOSO, AND E. B. REGITANO. 1989. Citrus leprosis mite Brevipal- pus phoenicis (Geijskes, 1939): Field test of chemical control. Anais da Escola Sup. Agric. Luiz de Queiroz 46: 473-483. MUMA, M. H. 1961. Mites associated with citrus in Florida. Florida Agric. Exp. Sta. Bull. 640. PRATT, R. M. 1958. Florida guide to citrus insects, diseases and nutritional disorders in color. Agric. Exp. Sta., Gainesville, FL. SAS INSTITUTE. 1991. SAS language and procedures: Usage 2, Version 6, First ed. SAS Institute, Cary, NC. SUDOI, V. 1990. Evaluation of different acaricides for control of red crevice mite Brevipalpus phoenicis Geijskes (Acari: Tenuipalpidae) infesting tea. Trop. Pest Management 36: 349-352.

272 Florida Entomologist 77(2) June, 1994

DEVELOPMENT OF NESTS AND COMPOSITION OF COLONIES OF NASUTITERMES NIGRICEPS (ISOPTERA: TERMITIDAE) IN THE MANGROVES OF JAMAICA

P. A. CLARKE AND ERIC GARRAWAY Department of Zoology, The University of the West Indies, Mona, Kingston 7, Jamaica West Indies

ABSTRACT

Colonies of Nasutitermes nigriceps (Haldeman) inhabiting the island of Jamaica were extracted from their arboreal carton nests by a two-stage method in which the termites were dislodged from their galleries and separated from nest debris by ﬂota- tion in water. Recovery of termites by this process was 97.7%. No incipient nests were observed and all small nests collected consisted of both a wooden region, which housed the reproductives, and an external carton nest. Except for the soldier caste, the proportion of sterile castes was similar in both regions of the nest. The size of small nests increased with little changes in the total colony size. However, the number of termites in the external carton nest increased with size of colony. Approximately 50% of the large nests contained reproductives. These colonies were consistently monogynous and monandrous. Caste composition of N. nigriceps colonies was comparable at each sample site and in nests of both reproductive status. However, large nests had a smaller proportion of immature termites, but similar proportion of soldiers. The average percentage of workers in small nests was proportional to that of large nests with reproductives, but less than large nests without reproductives. Key Words: Nasutitermes, termites nest, colony composition.

RESUMEN

Las colonias de Nasutitermes nigriceps (Haldeman) que habitan la isla de Ja- maica fueron extraidas de sus nidos arboreos por un método de dos pasos mediante el cual las termitas se separaron de sus galerías y se separaron del ripio usando un metodo de flotación en agua. El porcentaje de termitas recuperadas fué del 97.7%. No se observaron nidos incipientes y todos los nidos pequeños consisitieron de una parte leñosa , en la cual se encontraron los estados reproductores, y un nido ex- terno de cartón. A excepción de la casta de soldados, la proporción de castas esteriles fué similar en ambas partes del nido. El incremento del tamaño de los nidos pequeños no significó un gran cambio en el tamaño de la colonia. El 50 % de los nidos grandes contenían estados reproductores. Estas colonias eran consistentemente monoginas y monandras. La composición de ambos estados repro- ductivos de las colonias de N. nigriceps fué comparable en cada sitio muestreado y en cada nido. Sinembargo, nidos grandes tuvieron una proporción menor de estados juveniles de termitas, pero una proporción similar de soldados. El porcentaje promedio de obreras en los nidos pequeños fue proporcional al de los nidos grandes con estados reproductores, pero los nidos pequeños tuvieron un porcentaje menor de obreras que los nidos grandes sin estados reproductores. Las colonias de Nasutitermes nigriceps (Haldeman) que habitan la isla de Jamaica fueron extraidas de sus nidos arboreos por un método de dos pasos mediante el cual las termitas se separaron de sus gal- erías y se separaron del ripio usando un metodo de flotación en agua. El porcentaje de termitas recuperadas fué del 97.7%.

———————————— This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.uﬂ.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

Clarke & Garraway: Nasutitermes Nests and Composition 273

The Caribbean island of Jamaica is the third largest in the West Indies and has 16 documented species of termites belonging to the families Kalotermitidae, Rhino- termitidae and Termitidae (Snyder 1956). Advanced species of the family Termitidae are represented by Termes hispaniolae (Banks), Nasutitermes nigriceps (Haldeman), Nasutitermes costalis (Holmgren), Nasutitermes hubbardi Banks. N. nigriceps is the most common and prominent species of arboreal termite on the island with populations distributed in both natural forests and human-developed habitats. Advanced species of termites enhance their survival by evolving specialized castes protected in tough and complex nests. Previous studies of the composition of Nasutitermes colonies have been documented in N. rippertii (Rambur) (Krecek 1970), N. costalis (Krecek 1970, Clarke 1990), N. corniger (Motschulsky) and N. ephratae (Holmgren) (Thorne 1985), N. exitiosus (Hill) (Watson & Abbey 1987). Unfortunately, the data are not comparable because of differences in extraction methods and accuracy of results. The problem of ascertaining the composition and size of N. nigriceps colonies was surmounted in this study by the use of a simple and replicative extraction process, which yields almost full recovery of termites in nests. The density of nests and growth pattern of colonies, including the proportion of castes at various stages of development, were examined.

MATERIALS AND METHODS

The study focused on populations of N. nigriceps inhabiting mangrove forests because this environment offered virtually undisturbed, accessible nests that could provide an accurate description of colony and nest development. The three study sites selected were Port Royal, Hellshire and the North Coast. Four species of mangroves occurred in these areas. In order of abundance these are Rhizophora mangle L. (red mangrove), Avicennia germinans L. (black mangrove), Laguncularia racemosa Gaertn (white mangrove) and Conocarpus erectus L. (buttonwood mangrove). Before assessment of colonies commenced, the density of nests at each site was determined by counting the number of live nests in 6 randomly chosen 30 m x 30 m quadrats. Vi- ability of nests was visibly assessed by creating holes in the upper, mid and lower regions of nests and observing the emergence or non-emergence of termites. Between June 1987 and May 1988, five nests per month were randomly selected from each site. Each nest was half an ellipsoid in shape (Fig. 1) and the longest perimeter, height and diameter of base were measured before removal of nest from the supporting branch. These measurements were used to calculate the volume of the nest. Both the carton and wooden sections of small nests were removed for analysis, while those nests consisting solely of a carton structure were dislodged from the supporting branch by repeatedly hitting the branch with a hatchet. Each nest was placed in a plastic bag and transported to the laboratory where it was first subdivided and fumigated with carbon dioxide in reinforced plastic bags for 10-15 min. During stage one of the extraction process each piece of subdivided nest was further broken into small pieces and shaken by hand to dislodge the termites from the galleries. The termites and nest debris resulting from shaking all the pieces of a whole nest were collected in a tray and the empty pieces of nest from which the termites had been removed were discarded. Termites were separated from the nest debris during stage two of the extraction process by flotation in water. The total mixture of termites and nest debris derived from a single nest was submerged in water and allowed to stand for approximately 30 min. The nest material rapidly becomes saturated and sinks; then the floating termites can be easily removed. For a clearer separation, small volumes of the total sus- pension were successively removed, stirred in water and allowed to stand for 10 min. The water with the floating termites was decanted into a double sieve with upper and lower mesh sizes of 0.05 mm and 0.025 mm, respectively. The remaining debris

274 Florida Entomologist 77(2) June, 1994

Fig. 1. Structure of nest. in the bucket was rewashed with water to ensure complete removal of termites. The water was drained from the termites and the total amount of termites measured. Three sub-samples of 5 ml each were removed and the number of each caste counted. The total colony size and caste composition were calculated. The efficiency of the two-stage extraction method was determined by processing 10 nests and assessing the amount of termites remaining in a) the pieces of nests to be discarded after shaking the termites from the galleries and b) the waste residue in the bucket after flotation. Unlike the powdery residue in the bucket after stage two, the pieces of nests to be discarded after shaking were crumbled before flotation. Complete retrieval of termites was determined by visual observation. The mean percentages and standard deviation of sterile castes were calculated from Arcsine transformed data (Sokal & Rohl 1981, Zar 1984). Only re-transformed data are quoted in this paper; consequently, for some results the total percentage of castes per nest are not equal to 100. Analysis of variance and Student's "t" test were used for comparisons of means.

RESULTS

The density of N. nigriceps nests varied in each sample site (Table 1). Port Royal mangroves supported the highest number of live termite nests and, contrary to the other sites, some nests were situated in proximity to each other. Consequently, during the monthly sampling, the number of nests collected from Port Royal was significantly greater than Hellshire and the North Coast. Incipient nests were not observed at the three sites and only a few small nests (N = 12) were found in the mangroves of Port Royal. These small nests occurred in proximity to each other while larger nests were widely separated (Fig. 2; Spearman Rank test r = -0.8, P > 0.05). Each small nest consisted of an interconnecting wooden and an external carton nest. The wooden nest was ramiﬁed with galleries and some areas were lined with

Clarke & Garraway: Nasutitermes Nests and Composition 275

TABLE 1. DENSITY OF NESTS IN THE THREE STUDY SITES. No. of Live Nests1 Quadrat Port Royal Hellshire North Coast 1 0 (1) 0 (1) 1 2 2 (1) 1 0 (1) 3 3 0 (1) 1 4 2 0 (1) 1 5 511 6 3 (1) 1 1 x 2 0.5 1

1( ) No. of dead nests.

stercoral carton. The perimeter of the friable, external carton nest ranged from 10-35 cm. The number of termites in the external carton nest increased with colony size (Fig. 3). Large nests with perimeters 34-130 cm (N = 101) consisted solely of an external carton nest. The colony size of these large nests increased proportionally with nest size (r = 0.5, P > 0.001), while in smaller nests there was an increase in the size of nests with no corresponding increase in the number of termites (Fig. 4). N. nigriceps nests were enlarged during the island's bimodal rainy season that peaks in May and October. The direction of growth of small nests was multilateral, but mainly negatively geotropic in large nests. The latter pattern of nest expansion was consistent in large nests that had fallen from branches. The lower and older region of large nests with perimeter greater than 87 cm were devoid of termites. The

Fig. 2. The relationship between the density (No. per 30m2) and perimeter of nests.

276 Florida Entomologist 77(2) June, 1994

Fig. 3. The effect of colony growth on the distribution of termites in the wooden and carton regions of nests. walls were paler in color and separated easily between the fingers. These vacant galleries were invaded by inquilines. The shaking-flotation extraction method produced a clean sample of sterile termites devoid of nest debris. The mean number of termites per 5 ml ranged from 941- 1195 (Table 2). The average recovery of termites by shaking and flotation was 98.3% and 97.6%, respectively (Table 2). Total recovery of termites utilizing the two-stage method was 97.7%. Colonies of N. nigriceps consisted of the typical castes found in termite societies. All small nests contained a pair of reproductives in the wooden nest region. Approximately half of the large nests had a pair of reproductives protected by a fortified royal chamber embedded within the core of the nest. The remaining nests of similar sizes did not have reproductives and no royal chambers were found. Imma-

Clarke & Garraway: Nasutitermes Nests and Composition 277

Fig. 4. The relationship between the size of colonies and volume of nests.

ture termites and workers of small nests were equally distributed in the wooden and carton regions (Table 3; P > 0.05), while soldiers occurred in large proportions in the carton nest (P < 0.01). Although the proportion of sterile castes was highly varied in

TABLE 2. RECOVERY OF N. NIGRICEPS FROM NESTS USING A TWO-STAGE EXTRACTION METHOD. Percentage of Termites Recovered Mean No. of Termites per Nest No. Shaking Washing Total 5ml 1 97.2 93.7 95.5 1 195 2 99.2 98.6 98.9 964 3 99.0 91.3 95.2 1 066 4 98.5 95.8 97.2 1 046 5 98.6 98.6 98.6 941 6 92.2 98.5 95.4 987 7 92.6 97.4 95.0 1 083 8 99.1 98.9 99.0 1 034 9 98.8 99.3 99.1 1 001 10 99.9 99.2 99.6 1 146 Mean1 98.3 97.6 97.7 SD (min, max) 2.9, 1.5 2.9, 1.7 2.2, 1.5

1X and SD were calculated from Arcsine transformed data. 278 Florida Entomologist 77(2) June, 1994

TABLE 3. THE MEAN PERCENTAGE (CALCULATED FROM ARCSINE TRANSFORMED DATA) OF CASTES IN TWELVE SMALL NESTS OF N. NIGRICEPS. Section of Nest1 Castes Wooden Region Carton Region Larvae 20.9 (8.1; 37.9) 21.0 (4.7; 44.7) Presoldiers 1.3 (0.4; 2.7) 1.2 (0.3; 2.6) Soldiers 12.4 (7.6; 18.1) 17.1 (5.9; 32.4) Workers 63.3 (45.1; 79.9) 55.2 (45.1; 79.7)

1Minimum and maximum SD in ( ). large nests, the composition of castes was not inﬂuenced by geographical location and presence of reproductives (Table 4). Small nests from Port Royal had a higher percentage of larvae and presoldiers (immature individuals) than large nests (P < 0.01). The proportion of workers in small nests was similar to that of large nests with reproductives (P < 0.01) but less than that of large nests without reproductives (P > 0.05). Soldiers were equally represented in colonies at different stages of nest development and reproductive status.

DISCUSSION

A N. nigriceps colony is initiated in a tree cavity by alates, and wooden galleries are created to house the expanding colony. At a later stage of nest development an external carton nest is built with walls composed primarily of stercoral material (partially digested wood). Initially, the nest material is malleable, but hardens with age. Very little information is known about the dynamics of incipient colonies in the ﬁeld because of the clandestine nature of the termites and the absence of detectable external diagnostic features of nests. However, N. nigriceps colonies inhabiting small nests with both wooden and external carton regions, increased carton nest space

TABLE 4. THE MEAN PERCENTAGES (CALCULATED FROM ARCSINE TRANSFORMED DATA) OF CASTES IN LARGE NESTS OF N. NIGRICEPS. Sites1 Castes Port Royal Hellshire N.W. Coast Reproductives Present Larvae 18.8 (11.1,25.9) 20.6 (16.6,25.5) 19.5 (19.4,19.6) Presoldiers 1.8 (0.6,3.7) 1.6 (1.6,25.5) 1.1 (0.9,1.6) Soldiers 14.5 (10.1,43.7) 14.5 (11.3,18.0) 13.7 (10.5,16.9) Workers 80.7 (53.7,73.9) 78.8 (56.7,68.6) 82.4 (59.3,71.1) N 3088

Reproductives Absent Larvae 16.8 (9.5,27.6) 23.7 (13.1,36.6) 33.4 (9.3,36.2) Presoldiers 1.7 (0.4,3.3) 1.4 (0.3,3.0) 1.3 (0.0,2.0) Soldiers 13.1 (9.1,14.3) 13.6 (10.2,17.0) 13.1 (10.7,17.3) Workers 84.7 (55.9,78.0) 75.5 (49.1: 71.4) 76.4 (49.5:72.3) 30 12 13

1Minimum and maximum SD in ( ). Clarke & Garraway: Nasutitermes Nests and Composition 279 with limited increase in total colony size. Such activity not only allows rapid development of a nest that offers better protection from biotic and abiotic factors, but also creates an environment with a more stable and optimum micro-climate. The original wooden nest is abandoned and the colony continues development in the carton nest. Colony composition of N. nigriceps varies with the stage of nest development. How- ever, the proportion of each caste seems to stabilize in large colonies at different locations and is not influnced by proportional increase in colony and nest size. Thus, the data presented in this paper may be applied to populations throughout Jamaica. Mature colonies accommodate the increasing numbers of termites by seasonal expansion of nests. During the rainy season, N. nigriceps workers build in a negative geotropic direction. Migration of termites within nests also follow a similar pattern so that termites are continuously associated with fairly new stercoral walls. This inter-relationship may be linked to the homeostatic properties of stercoral material. A review of Noirot (1970) showed the positive influence of stercoral walls on temperature and humidity within nests. The homeostatic nature of stercoral walls of N. nigriceps nests may decline with age as seen in its physical parameters such as texture and color. Further research is required for confirmation. N. nigriceps colonies with reproductives were consistently monogynous and monandrous, but approximately half the nests were devoid of reproductives. The latter may be either calies (subsidiary nests) or large nests which lost their reproductives. Similarity in size of both types of nests, comparable caste composition, absence of a royal chamber in queenless nests and distance between nests, suggest that these large nests without reproductives are calies. Calies have never been recorded in N. nigriceps, but were documented in some members of the genus; N. corniger (Thorne 1982), N. costalis and N. polygynous (Roisin and Pasteels 1986). While it is still not clear what induces the development of calies, it has been suggested that they are constructed in response to adverse conditions created by increase colony size (Holt & Easey 1985; Adams & Levings 1987). A hardened nest inhabited by a large colony hinders accurate determination of the size and composition of the colony. The use of shaking and flotation as methods of collecting insects is not a novelty (Southwood 1978), however, the two-stage method described here has never been utilized to extract termites of the genus Nasutitermes. This method is comparable with that used by Darlington (1984) and Thorne (1985). Thorne (1985) removed N. corniger and N. ephratae (after inactivation by freezing) by a similar shaking method, but the termites were not separated from the nest debris before subsampling thus, resulting in a variation of 400-900 termites per 5 ml subsample. This highlighted the need for a second stage in the extraction method that would provide a cleaner separation and ultimately increase precision. Darling- ton's (1984) study of Macrotermes spp. found flotation useful in separating termites from the rubble generated from the dissection of their huge nests, hence developed a special flotation tank. The two stage method used in this study allows rapid collection of accurate data with the use of simple equipment such as buckets, trays and a portable 10 lb cylinder of carbon dioxide.

ACKNOWLEDGMENTS

The research was funded by The University of the West Indies. We thank Dionne Miller and Johanna P.E.C. Darlington for useful discussions.

REFERENCES CITED

ADAMS, E. S., AND S. C. LEVINGS. 1987. Teritory size and population limits in mangroves termites. J. Anim. Ecol. 56: 1069-1081. 280 Florida Entomologist 77(2) June, 1994

CLARKE, P. 1990. Caste ratios in parent nests and calies of Nasutitermes costalis (Holmgren). U.W.I. Biospectrum 2: 15-16. DARLINGTON, J. P. E. C. 1984. A method for sampling the population of large termite nests. Ann. Appl. Biol. 104: 427-436. HOLT, J. A., AND F. J. EASEY. 1985. Polycalic colonies of some mound-building termites (Isoptera: Termitidae) in Northeastern Australia. Insectes Sociaux, Paris 32: 61-69. KRECEK, J. 1970. Nest structure, humidity and colony composition of two species of Nasutitermes in Cuba (Isoptera: Termitidae). Act. Entomol. Bohemoslauoca, 67: 310-317. NOIROT, C. 1970. The nests of termites, in K. Krishna and F. M. Weesner [eds.], Biol- ogy of Termites. Vol. II., Academic Press, New York, London. 693 pp. . ROISIN, Y., AND J. M. PASTEELS. 1986. Reproductive mechanisms in termites: poly- calism and polygyny in Nasutitermes polygynous and Nasutitermes costalis. Insectes Sociaux, Paris 33: 149-167. SNYDER, T. E. 1956. Termites of the West Indies, the Bahamas and Bermuda. J. Agric. of the University of Puerto Rico 60: 189-201. SOKAL, R. R., AND F. J. ROHLF. 1981. Biometry. W. H. Freeman and Company, New York. 859 pp. SOUTHWOOD, T. R. E. 1978. Ecological methods. Chapman and Hall. New York, Lon- don. 379 pp. THORNE, B. L. 1985. Numerical and biomass caste proportions of colonies of termites Nasutitermes corniger and Nasutitermes ephratae (Isoptera: Termitidae). In- sectes Sociaux, Paris 32: 411-426. WATSON, J. A. L., AND H. M. ABBEY. 1987. Maternal determination of reproductives vs sterile castes in Nasutitermes exitiosus (Hill) (Isoptera: Termitidae). In- sectes Sociaux, Paris, 34: 291-297. ZAR, J. H. 1984. Biostatistical Analysis. Prentice Hall, New Jersey. pp. 718.

Scientiﬁc Notes 281

APPLICATION METHODS FOR ENTOMOPATHOGENIC NEMATODES (RHABDITIDA: HETERORHABDITIDAE): AQUEOUS SUSPENSIONS VERSUS INFECTED CADAVERS

RICHARD K. JANSSON AND SCOTT H. LECRONE Tropical Research and Education Center Institute of Food and Agricultural Sciences University of Florida Homestead, FL 33031

Entomopathogenic nematodes have potential as biological control agents of many soil insect pests. These nematodes have a broad host range, and they are highly vir- ulent with high reproductive rates. They have the ability to seek out and quickly kill hosts (even cryptic hosts) within 24-48 h, and they are safe to vertebrate and other nontarget organisms (Gaugler 1981, Kaya 1985). In addition, these nematodes are easy to apply using a variety of methods ranging from aqueous suspensions applied by hand to those made with standard sprayers (up to 70.4 kg/cm2) and irrigation systems (Kaya 1985, Georgis 1990). Entomopathogenic nematodes have also been shown to have potential for controlling field populations of the sweetpotato weevil, Cylas formicarius (Fabricius) (Jans- son et al. 1990, 1991, 1993). Heterorhabditid nematodes were superior to steinernematids at reducing weevil populations and their concomitant damage to storage roots (Jansson et al. 1990, 1993). Welch & Briand (1960) found that aqueous applications of Steinernema carpocapsae (Weiser) DD136 strain, and those made using infected greater wax moth, Galleria mellonella (L.), cadavers, were equally effective at controlling cabbage root maggot, Hylemya brassicae (Bouché). More recently, Jansson et al. (1993) showed that applications of G. mellonella cadavers infected with heterorhabditid nematodes were efficacious for controlling field populations of C. formicarius and their concomitant damage; however, nematode applications via infected cadavers were not compared with those made via aqueous suspensions. The present study compared efficacy and persistence of a heterorhabditid nematode against C. formicarius when applied via aqueous suspensions with those made using infected G. mellonella cadavers. The experiment was conducted in a Krome, very gravelly loam soil at the Tropical Research and Education Center in Homestead. Sweet potato, Ipomoea batatas (L.) Lam. cv. Jewel, transplants were hand planted 20 cm apart on raised beds with cen- ters spaced 1.9 m apart on 5 June 1991. All production practices were similar to those described previously (Jansson et al. 1990, 1991, 1993). No herbicides or fungi- cides were applied to the experimental plot. Plants were drip irrigated 4 h per day using a drip turbo T-tape-irrigation system (model 40) (5.0 liter per m per h) from shortly before planting until the experiment terminated. A research plot (0.2 ha) was subdivided and arranged into a randomized complete block design with four replications. Treatment plots were three beds by 15.2 m long. A 3-m buffer of nontreated plants separated replicates. Treatments evaluated were: a single aqueous application of Heterorhabditis bacteriophora Poinar HP88 strain (4.9 billion infective juveniles per ha) made on 23 July 1991, a single application of 5- and 10-day-old G. mellonella cadavers (83,700 per ha) infected with H. bacteriophora HP88 strain made on 26 July 1991, and nontreated plants. Jansson et al. (1991) showed that a single application of H. bacteriophora HP88 strain was efficacious for controlling populations of C. formicarius. Potential rates of infective juveniles applied in the cadaver treatments were 6.4 to 18.3 billion infective juveniles per ha based on mean numbers of infective juveniles produced per cadaver of between

282 Florida Entomologist 77(2) June, 1994

76,260 and 219,181 (Jansson et al. 1993). Inoculation procedures were similar to those described previously (Jansson et al. 1993). Two 3-m sections were dug from the middle bed of each treatment plot on 19-20 December 1991. Storage roots were divided into three size categories (small, medium, and jumbo) and each individual root was then visually inspected for weevil damage and rated on a scale from 1 to 6 (Jansson et al. 1990): 1, no weevil damage, no feeding or oviposition scars, no adult exit holes; 6, severe weevil damage, > 6 adult exit holes per root. The percentage of root biomass in each damage category and the mean damage index were determined. The percentage of marketable roots was determined by calculating the percentage of medium-sized roots with a rating ≤ 2 (Jan- sson et al. 1990). Soil was assayed for native nematode populations once [19 July 1991, 12 soil samples (720-1045 g each) per plot] before applications were made. Levels of recovery of HP88 nematodes were determined on three dates after application (19 September, 5 December 1991 and 23 July 1992; 3 soil samples per treatment plot) using methods described previously (Jansson et al. 1991, 1993). Soil was transferred to separate plastic cups (1 liter) and ten late-instar G. mellonella were placed in the bottom of each cup, covered with soil (Bedding & Akhurst 1975) and stored for 10-14 days in the dark at 24±3°C after which cadavers were examined for nematode infection (Jan- sson et al. 1991, 1993). Percentages of root weight in various damage categories were transformed to the arcsine of the square root and analyzed by least squares analysis of variance (Zar 1984). Total numbers of infected G. mellonella larvae were compared among treatments by X2 analysis (Steel & Torrie 1980). Percentages of roots that had no weevil damage (rating = 1), slight weevil damage (rating = 2), or little or no damage (rating = 1 or 2) did not differ (F ≥ 0.9, df = 3,9, P > 0.05) among treatments (Table 1). Percentages of root weight in these three damage categories tended to be higher on nematode-treated plants than on nontreated plants, although not signiﬁcantly. These data suggest that aqueous applications of nematodes were comparable in efﬁcacy to those made using infected cadavers of G. mellonella that were either 5 or 10 days old. Few soil samples (0.7%, n = 388) and fewer G. mellonella larvae (0.1%, n = 3,880) were positive for entomopathogenic nematodes before applications were made. After

TABLE 1. PERCENTAGES OF STORAGE ROOT WEIGHT WITH NO WEEVIL DAMAGE, SLIGHT WEEVIL DAMAGE, AND NONE OR SLIGHT DAMAGE, ON SWEET POTATOES TREATED WITH AQUEOUS OR CADAVER APPLICATIONS OF H. BACTERIO- PHORA HP88 STRAIN AND ON NONTREATED PLANTS. % Root Weight 1 Treatment No Damage, Slight Damage, Little/No Damage, Rate/ha2 Rating = 1 Rating = 2 Rating = 1 or 2 Aqueous, 4.9 B/ha 50.6±3.6 28.3±1.8 79.0±3.1 Cadaver, 5-day-old, 83,700/ha 47.9±6.9 31.1±3.6 79.0±3.7 Cadaver, 10-day-old, 83,700/ha 51.5±3.7 28.8±1.6 80.4±3.6 Nontreated 34.6±4.8 32.1±5.4 66.7±4.2

1Means within a column did not differ (P > 0.05) by least squares analysis of variance (Zar 1984). 2Aqueous applications are in terms of infective juveniles per ha; cadaver treatments are numbers of cadavers per ha.

Scientiﬁc Notes 283

Fig. 1. Total numbers of G. mellonella larvae infected with H. bacteriophora HP88 strain in sweet potato plots treated with aqueous suspensions of HP88 or cadavers of G. mellonella infected with HP88 and in nontreated plots (n= 120 larvae per treatment). applications, high levels of recovery of nematodes were found on the ﬁrst two sample dates (55-58 and 132-135 days after application) (Fig. 1). Little recovery was found on the last sample date (362-365 days after application) approximately 7 months after the experiment terminated. Recovery of nematodes did not differ (X2 ≤ 8.8, df = 2, P > 0.05) among the three nematode treatments, but was considerably higher in nematode-treated plots than in nontreated plots (Fig. 1). Levels of recovery concur with our previous experiments in which either aqueous suspensions or cadaver applications were used (Jansson et al. 1991, 1993). The data show that aqueous suspensions of HP88 nematodes were as effective as applications of G. mellonella cadavers infected with HP88 for controlling damage by sweetpotato weevil to storage roots. Also, we found that persistence of nematodes was similar for the two of methods of application. Application of infected cadavers for introducing nematodes was shown to have merit in a previous report (Jansson et al. 1993). Applications of nematodes via infected insect hosts may have potential for in- tegrating into developing countries because its simple and requires no special equipment and no water at the time of application. However, it does require a constant and large supply of susceptible hosts and considerable space and labor. We encourage re- searchers to explore the possibility of using this approach in a variety of insect host/ crop systems. We thank E. Murray and R. Lance for assistance with data collection and the anonymous reviewers for their suggestions. This research was supported by U.S.D.A., C.S.R.S., Tropical/Subtropical Agriculture Program, Grant Nos. 88-34135-3564 and 91-34135-6134 (to R.K.J.) managed by the Caribbean Basin Ad- ministrative Group (CBAG). The senior author is currently Senior Research Fellow, Merck Research Laboratories, P. O. Box 450, Hillsborough Rd., Three Bridges, NJ 08887-0450. This is Florida Agricultural Experiment Station Journal Series No. R-03090.

284 Florida Entomologist 77(2) June, 1994

SUMMARY

Aqueous suspensions of HP88 nematodes were as effective as applications of Gal- leria mellonella cadavers infected with HP88 for controlling damage by sweetpotato weevil to storage roots. Persistence of nematodes was similar for the two types of application.

REFERENCES CITED

GAUGLER, R. 1981. Biological control potential of neoaplectanid nematodes. J. Nem- atol. 13: 241-249. GEORGIS, R. 1990. Formulation and application technology, p. 173-191 in R. Gaugler and H. K. Kaya [eds.], Entomopathogenic nematodes in biological control. CRC Press, Boca Raton, Florida, 356 p. JANSSON, R. K., S. H. LECRONE, AND R. GAUGLER. 1991. Comparison of single and multiple releases of Heterorhabditis bacteriophora Poinar (Nematoda: Heter- orhabditidae) for control of Cylas formicarius (Fabricius) (Coleoptera: Apion- idae). Biol. Control 1: 320-328. JANSSON, R. K., S. H. LECRONE, AND R. GAUGLER. 1993. Field efﬁcacy and persistence of entomopathogenic nematodes (Rhabditida: Steinernematidae, Heter- orhabditidae) for control of sweetpotato weevil (Coleoptera: Apionidae) in southern Florida. J. Econ. Entomol. 86: 1055-1063. JANSSON, R. K., S. H. LECRONE, R. R. GAUGLER, AND G. C. SMART, JR. 1990. Potential of entomopathogenic nematodes as biological control agents of the sweetpotato weevil (Coleoptera: Curculionidae). J. Econ. Entomol. 83: 1818-1826. KAYA, H. K. 1985. Entomogenous nematodes for insect control in IPM systems, p. 283-302 in M. A. Hoy and D. C. Herzog [eds.], Biological control in agricultural IPM systems. Academic Press, New York. STEEL, R. G., AND J. H. TORRIE. 1980. Principles and procedures of statistics. McGraw-Hill, New York, 633 p. WELCH, H. E., AND L. J. BRIAND. 1960. Field experiment on the use of a nematode for control of vegetable crop insects. Proc. Entomol. Soc. Ontario 91: 197-202. ZAR, J. H. 1984. Biostatistical analysis. Prentice-Hall, Englewood Cliffs, New Jersey, 718 p.

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284 Florida Entomologist 77(2) June, 1994

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EFFECT OF TILL AND NO-TILL SOYBEAN CULTIVATION ON DYNAMICS OF ENTOMOPATHOGENIC FUNGI IN THE SOIL

D. R. SOSA-GOMEZ AND F. MOSCARDI EMBRAPA - Centro Nacional de Pesquisa de Soja, Cx P 1061. Londrina, PR - 86001-970. Brazil.

Several species of phytophagous insects are attacked by entomopathogenic fungi of the Moniliacea family. In soybean agroecosystems, the most prevalent fungi are Nomuraea rileyi (Farlow) Samson which infect caterpillars, and Beauveria bassiana (Balsamo) Vuill. which attacks chrysomelids such as Diabrotica speciosa (Germar), Cerotoma arcuata (Olivier), Colaspis spp. and curculionids, such as Aracanthus spp. Other fungal species of lesser prevalence associated with soybean insects include Metarhizium anisopliae (Metsch.) Sorok., which is found on Phyllophaga cuyabana (Moser) and other Scarabaeidae; Paecilomyces fumosoroseus (Wize) Brown & Smith which cause epizootics in populations of Lagria villosa Fabr. (Coleoptera: Lagriidae); and another species of Paecilomyces, probably tenuipes, which occurs at low levels in the wet season on Anticarsia gemmatalis Hubner, Chrysodeixis includens (Walker)

Shelly: Methyl eugenol consumption by Bactrocera dorsalis 285 and Rachiplusia nu (Guenée) (D. Sosa-Gómez unpublished). The prevalence of these entomopathogenic fungi are affected by various abiotic and biotic factors including cultural practices (Gaugler et al. 1989; Sosa-Gómez & Moscardi 1992). The objective of this paper was to study the effect of till and no-till cultivation practices on density dynamics of entomopathogenic fungi in the soil. The experiment was carried out in two contiguous areas of 22 x 312 m each, in Londrina, state of Parana, Brazil, one under no-till cultivation and another under conventional tillage. In the area under no-till, the soil was not cultivated during the entire year. Double-croppings of soybean and wheat had been planted on these areas since 1985. The soil type was classified as oxisols (soil taxonomy of Brazil = latossolo roxo distrófico epieutrófico). Along the length of the two areas, nine paired plots (50 m2) were delimited in the 1989/90 season and eight paired plots (50 m2) in the 1990/ 91 season. In each paired plot, eight soil core subsamples, 2 cm diam and about 2 cm deep, were taken every two weeks from the soil surface from randomly selected sites (1 m2). The majority of fungus inocula usually concentrate in this soil layer (Ignoffo et al. 1977; Storey et al. 1989). The subsamples from each plot were combined and mixed thoroughly in the laboratory. One-gram aliquots were taken per sample and processed for colony forming units (CFUs) growing on selective medium (Chase et al. 1986). The number of CFUs per gram of dry soil was analyzed by the t-test procedure of the Statistical Analysis System (SAS Institute 1985), so as to compare mean responses between CFUs from tilled and untilled paired plots. Significant differences (P < 0.05) in the number of CFUs recovered from the two areas occurred at practically all sampling dates (Fig. 1). The predominant species among the observed entomopathogenic fungi was B. bassiana reaching about 1.9 x 105 CFUs per g of soil on February 2, 1990 (Fig. 1a) and 3.3 x 104 CFUs per g of soil on January 17, 1991 (Fig. 1b). The prevalence of B. bassiana explains the frequent incidence of this fungus on populations of chrysomelids and pentatomids as reported by Moscardi et al. (1985) and Daoust & Pereira (1986). M. anisopliae reached the maximum level on January 19, 1990 with 1.2 x 104 CFUs per g of soil (Fig. 1c), and 1.1 x 104 CFUs/g soil in 1991 (Fig. 1d). Paecilomyces spp. reached a peak incidence of 1.9 x 104 on December 14, 1989 (Fig. 1). Thus, in both years, the three fungus species occurred at higher levels in soils cultivated under the no-till system. The differences can be attributed to biotic and abiotic differences between the cultivation systems. Wierenga et al. (1982) mentioned that the surface temperature amplitudes are considerably larger in tilled soil than in non-tilled soil. Other factors, such as the soil ca- pacity to retain water, higher organic matter, and lower temperature of soils under minimum tillage (Vieira 1981) may have contributed to the differential prevalence of fungi under the two cultivation systems studied. Gaugler et al. (1989) observed that application of B. bassiana followed by tillage was very important in achieving enhanced fungal persistence in the soil. This was attributed to the incorporation of conidia into the soil. In the same way, no-tillage practices can enhance persistence of entomopathogenic fungi in the surface layer of the soil, as found in the present work. More research is needed to evaluate the effect of the greater persistence and prevalence of fungi in no-till systems on natural populations of soybean pests as compared to conventional tillage systems. This information will be important for devising cultural practices aimed at augmenting the natural occurrence of entomopathogenic fungi on soybean insects and, consequently, increasing the contribution of natural pest mortality in soybean agroecosystems. We thank Ivanilda Soldorio and Jairo da Silva for technical assistance.

SUMMARY

Cultural practices inﬂuence propagule densities of entomopathogenic fungi in agroecosystems. An experiment was conducted in Brazil to evaluate the prevalence of

286 Florida Entomologist 77(2) June, 1994

Fig. 1. Effect of till and no-till farming on density of colony forming units (CFUs) of Beauveria bassiana (A, B), Metarhizium anisopliae (C, D) and Paecilomyces spp. (E, F) in two seasons. Means within each sample date with different letters are sig- niﬁcantly different. (P > 0.05; t-test).

Beauveria bassiana, Metarhizium anisopliae and Paecilomyces spp. in soil under no- till and conventional tillage systems. The no-till cultivation favored the prevalence of the three fungus species when compared with the conventional tillage system.

REFERENCES CITED

CHASE, A. R., L. S. OSBORNE, AND V. M. FERGUSON. 1986. Selective isolation of the entomopathogenic fungi Beauveria bassiana and Metarhizium anisopliae from an artiﬁcial potting medium. Florida Entomol. 69: 285-292. DAOUST, R. A., AND R. M. PEREIRA. 1986. Stability of entomopathogenic fungi Beau- veria bassiana and Metarhizium anisopliae on beetle-attracting tubers and cowpea foliage in Brazil. Environ. Entomol., 15: 1237-1243. GAUGLER, R., S. D. COSTA, AND J. LASHOMB. 1989. Stability and efﬁcacy of Beauveria bassiana soil inoculations. Environ. Entomol. 18: 412-417. IGNOFFO, C. M., C. GARCIA, D. L. HOSTETTER, AND R. E. PINNELL. 1977. Vertical movement of conidia of Nomuraea rileyi through sand and loam soils. Jour. Econ. Entomol. 70: 163-164. SAS INSTITUTE. 1985. Guide for personal computers. SAS Institute, Cary, N.C.

Shelly: Methyl eugenol consumption by Bactrocera dorsalis 287

MOSCARDI, F., B. S. CORREA-FERREIRA, L. G. LEITE, AND C. E. O. ZAMATARO. 1985. Incidência estacional de fungos entomógenos sobre populações de percevejos pragas da soja, in Resultados de Pesquisa de Soja 1984/1985. p. 90. SOSA-GÓMEZ, D. R., AND F. MOSCARDI. 1992. Epizootiologia: chave dos problemas para o controle microbiano com fungos, in Simposio de Controle Biológico. Anais. p. 64- 69. CNPDA-EMBRAPA [ed.]. Aguas de Lindoia, 12-16/10/92. 312 p. STOREY, G. K., W. A. GARDNER, AND E. W. TOLLNER. 1989. Penetration and persistence of commercially formulated Beauveria bassiana conidia in soil of two tillage systems. Environ. Entomol. 18: 835-839. VIEIRA, M. J. 1981. Propriedades físicas do solo, in Plantio direto no estado do Paraná. Circular no 23. Fundação Instituto Agronômico do Paraná. p. 19-32. WIERENGA, R. J., D. R. NIELSEN, R. HORTON, AND B. HEIES. 1982. Tillage effects on soil temperature and thermal conductivity. In Predicting Tillage Effects on Soil Physical Properties and Processes. Asa Special Publication, 44. Ed. by American Society of Agronomy and Soil Science Society of America, Inc. Chap. 5. p. 69 - 90.

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Scientiﬁc Notes 287

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FRUIT OF MORRENIA ODORATA (ASCLEPIADACEAE) AS A HOST FOR THE PAPAYA FRUIT FLY, TOXOTRYPANA CURVICAUDA (DIPTERA: TEPHRITIDAE)

PETER J. LANDOLT U. S. Department of Agriculture, Agriculture Research Service 1700 S.W. 23rd. Dr., Gainesville, FL 32604

The papaya fruit fly, Toxotrypana curvicauda Gerstaecker, is a pest of papaya fruit, Carica papaya L. (Caricaceae), throughout much of the neotropics, including southern Florida in the United States. It had previously been considered restricted to papaya (Knab & Yothers 1914; Wolfenbarger & Walker 1974). However, it has been reported from mango in Florida (Butcher 1952) and from additional species of plants in Mexico (Castrejon-Ayala 1987). I report here that papaya fruit flies have been reared from field-collected fruit of Morrenia odorata Lindl., an asclepiad or milkweed, in Florida. Pods were collected from climbing vines of M. odorata on a palm tree in a suburban neighborhood of Sa- rasota during April 1993. These fruit were mature in size (6-7 cm long and 3-5 cm diam) but were still green when collected, and produced latex when broken at the stem. Fruit were held in a screened cage in a laboratory at 22°C and 45±5% RH. On 20 April 1993 thirty-four mature larvae emerged from one of two fruit collected in Sarasota 1 April 1993. Three additional fruit collected 8 April did not yield any fly larvae. The 34 larvae were placed in sterilized potting soil for pupation. Twelve male and 11 female adult papaya fruit flies emerged from 19 to 22 May 1993, 30-33 days after pupation. Papaya fruit flies have not been reported previously from this plant. Morrenia odorata was introduced into Florida, possibly from Argentina, and is widely distributed in the central area of the state. The utilization of such a widely oc- curring weed may conceivably promote a broader distribution of this pest insect in Florida, and make it easier for the papaya fruit fly to infest disjunct areas of papaya production. Papaya fruit flies have been reared from the fruit of other Asclepiadaceae, in Mexico. Gonolobus sorodius A. Gray is a host of T. curvicauda in central Mexico (Cas- trejon-Ayala 1987, Castrejon-Ayala & Camino-Lavin 1991). Also, Baker et al. (1944) reported talayote, or talayotillo (a vine in the milkweed family) as a host either of T. curvicauda or an undescribed species of Toxotrypana in northeast Mexico. This plant

288 Florida Entomologist 77(2) June, 1994 may be the same as the talayote of Martinez (1969), which is Gonolobus erianthus D.C., a climbing milkweed vine found in Morelos, Mexico. The milkweed genus Mor- renia is also a host of other species of Toxotrypana in Argentina (Blanchard 1959). Voucher specimens are placed in the Florida State Collection of Arthropods, Gainesville, Florida and in the National Museum of Natural History, Washington, D.C. The plant was identiﬁed by C. Artaud of the Florida Dept. Agriculture, Division of Plant Industry, Gainesville, Florida. I thank R. Knight for information on Asclepi- adaceae and A. L. Norrbom for information on Toxotrypana host records.

SUMMARY

Twenty three adult papaya fruit flies, Toxotrypana curvicauda Gerstaecker, were obtained from 34 mature larvae that emerged from field-collected fruit of the milkweed vine, Morrenia odorata Lindl. collected in Sarasota, Florida. Morrenia odorata appears to be an alternate host for the papaya fruit fly in Florida.

REFERENCES CITED

BAKER, A. C., W. E. STONE, C. C. PLUMMER, AND M. MCPHAIL. 1944. A review of studies on the Mexican fruit fly and related Mexican species. USDA Misc. Publ. No. 531, 155 pp. BLANCHARD, E. E. 1959. El genero Toxotrypana en la Republica Argentina (Diptera; Trypetidae). Acta Zool. Lilloana 17: 33-45. BUTCHER, F. G. 1952. The occurrence of papaya fruit fly in mango. Florida State Hort. Soc. Proc. 65: 196. CASTREJON-AYALA, F. 1987. Aspectos de biologia y habitos de Toxotrypana curvicauda Gerst. (Diptera: Tephritidae) en condiciones de laboratorio y su distribution en una plantacion de Carica papaya L. en Yuatepec, Mor. BS Thesis, Instituto Polictecnico Nacional, Mexico D.F., Mexico. 88 p. CASTREJON-AYALA, F., AND M. CAMINO-LAVIN. 1991. New host plant record for Tox- otrypana curvicauda (Diptera: Tephritidae). Florida Entomol. 74: 466. KNAB, F., AND W. W. YOTHERS. 1914. Papaya fruit fly. J. Agric. Res. 2: 447-453. MARTINEZ, M. 1969. Las Plantas Medicinales de Mexico. 5th Edition, Justo Sierra 52. Mexico 1, D.F. WOLFENBARGER, D. O., AND S. D. WALKER. 1974. Two major pest problems of papa- yas. Proc. Florida State Hort. Soc. 85: 384-385.

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288 Florida Entomologist 77(2) June, 1994

PESTICIDE RESISTANCE AND METABOLIC RATE IN GERMAN COCKROACHES (DICTYOPTERA: BLATTELLIDAE)

MARK E. HOSTETLER, JOHN F. ANDERSON, AND CARMINE A. LANCIANI Department of Zoology, University of Florida, 223 Bartram Hall, Gainesville, FL 32611, USA

Although physiological resistance mechanisms to most pesticides have developed in insects, costs may be associated with these mechanisms. For example, a reproductive cost is shown by resistant tobacco budworms, Heliothis virescens, which produce significantly fewer fertile eggs (Plapp et al. 1990), and a behavioral cost is shown by resistant Anopheles mosquitoes, which spend less time searching for hosts (Rowland 1987). An energetic cost may also be associated with resistance. For example, a common mechanism of pesticide detoxification is enzymatic breakdown of ingested or ab- This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.ufl.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

Scientific Notes 289 sorbed toxins (Soderland & Bloomquist 1990), a process that may increase energy use as detoxification enzymes are synthesized. To determine whether an energetic cost to pesticide resistance exists, we measured metabolic rates (O2 consumption) of resistant and susceptible German cockroaches, Blattella germanica (L.). German cockroaches used in experiments were obtained from 2 different strains maintained at the USDA-ARS Medical and Veterinary Entomology Research Labo- ratory in Gainesville, Florida, U.S.A. The Village Green strain is physiologically resistant to a variety of organophosphates and pyrethroids. Ancestors of this strain were captured from a residential apartment complex (Village Green Apartments) in Florida in 1987. The Orlando Normal strain is susceptible to a variety of organophosphates and pyrethroids. Ancestors of this strain were captured from a building in Or- lando, Florida, U.S.A., in the 1940s. Cockroaches were reared in 50 x 50 x 28 cm oblong metal tubs (temperature of 25-27oC) on Purina rat chow (Purina Mills Inc. P.O. Box 66812, St. Louis MO 63166-6812). Metabolic-rate experiments were conducted in Gainesville during October and November 1991 on male and female adults (gravid females without oothecae pro- truding) that had spent 0 to 2 weeks in the adult stage. Metabolic rates were measured with “closed system” metabolic chambers (Vleck, 1987) following the procedures described by Anderson et al. (1989) and Giesel et al. (1989). On the day of each experiment, 5 male and 5 female cockroaches from each strain were placed individually in 60 cm3 syringes. After the syringes were held for approximately 5 hours o in a lighted incubator at 26 C, each syringe was removed, and the change in O2 concentration was recorded and used to determine metabolic rate. Each cockroach was weighed to the nearest 0.01 mg immediately after removal from the incubator. Sep- arate trials were conducted on 4 different days, yielding data on 20 males and 20 females in both the Orlando Normal and Village Green strains. Because sex and weight may affect metabolic rate, analysis of covariance (AN- COVA) was used to determine the influence of strain on cockroach metabolic rates independent of the influence of sex and weight. Mean metabolic rates and weights of both strains are listed in Table 1. From the analysis of covariance, only gender (P = 0.0002) and weight (P = 0.0046), but not strain (P = 0.38), had significant effects on metabolic rates. Thus, after the effects of weight and strain were removed by the ANCOVA, females had a higher metabolic rate than did males. However, after the effects of weight and sex were removed by the ANCOVA, resistant individuals did not have a higher metabolic rate than did susceptible individuals. Resistant and susceptible strains had similar metabolic rates, and thus resistance may not have an energetic cost. Several explanations exist for the similar metabolic rates of the two strains. (1) The Village Green cockroaches may have physiological resistance mechanisms that do not require increased enzymatic activity, e.g., altered acetylcholinesterases and sodium channels (Soderland & Bloomquist 1990). (2) The Village Green cockroaches may have detoxification mechanisms that increase metabolic rates but require the presence of the pesticide to induce acceler- ated production of the detoxifying enzyme(s). In several studies, enzyme production-

µ TABLE 1. MEAN METABOLIC RATES ( L O2/HR + STANDARD ERROR) AND MEAN WEIGHTS (MG + STANDARD ERROR) OF BLATTELLA GERMANICA. Strain Sex N Metabolic Rate Weight Orlando M 20 34.9 + 1.2 43.3 + 0.9 Orlando F 20 64.3 + 4.1 84.3 + 4.4 VG M 20 31.6 + 1.1 48.3 + 0.8 VG F 20 73.4 + 4.7 94.8 + 4.1

290 Florida Entomologist 77(2) June, 1994 was found to increase only after exposure to the pesticide (Terriere 1983). (3) Village Green cockroaches may have higher metabolic rates from pesticidal pressures, but the Orlando Normal individuals may have higher than “normal” metabolic rates because they have been laboratory-reared for a much longer period of time. We thank the USDA-ARS, Medical & Veterinary Entomology Research Laboratory for providing cockroaches. This research was in part funded by the Clorox Company.

SUMMARY

Metabolic rates, based on oxygen consumption, were determined for two strains of German cockroaches, one resistant and one non-resistant. Both strains had similar metabolic rates suggesting that resistance may not have an energic cost.

REFERENCES CITED

ANDERSON, J. F., C. A. LANCIANI, AND J. T GIESEL. 1989. Diel cycles and measurement of metabolic rate in Drosophila. Comp. Biochem. Physiol. 94A: 269-271. GIESEL, J. T., C. A. LANCIANI, AND J. F. ANDERSON. 1989. Larval photoperiod and metabolic rate in Drosophila melanogaster. Florida Entomol. 72: 123-128. PLAPP, F. W., J. C. CAMPANHOLA, R. D. BAGWEL, AND B. F. MCCUTCHEN. 1990. Man- agement of pyrethroid-resistant tobacco budworms on cotton in the United States. pp. 237-260 in R. T. Roush and B. E. Tabashnik [eds.], Pesticide resistance in arthropods. Chapman and Hall, New York. ROWLAND, M. W. 1987. Fitness of insecticide resistance. Nature 327: 194. SODERLUND, D. M., AND J. R. BLOOMQUIST. 1990. Molecular mechanisms of insecticide resistance, pp. 58-96 in R. T. Roush and B. E. Tabashnik [eds.], Pesticide resistance in arthropods. Chapman and Hall, New York. TERRIERE, L. C. 1983. Enzyme induction, gene ampliﬁcation and insect resistance to insecticides, pp. 265-298 in G. P. Georghiou and T. Saito [eds.], Pest resistance to pesticides. Plenum Press, New York. VLECK, D. 1987. Measurement of O2 consumption, CO2 production, and water vapor production in a closed system. J. Appl. Physiol. 62: 2103-2106.

✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦✦

290 Florida Entomologist 77(2) June, 1994

MCPHAIL TRAP CAPTURES OF ANASTREPHA OBLIQUA AND ANASTREPHA LUDENS (DIPTERA: TEPHRITIDAE) IN RELATION TO TIME OF DAY

EDI A. MALO AND GEORGINA I. ZAPIEN Laboratorio de Biología y Ecología de Moscas de la Fruta, Centro de Investigaciones Ecológicas del Sureste, Carretera al Antiguo Aeropuerto Km. 2.5, Apartado postal 36, Tapachula, 30700, Chiapas, México

Anastrepha Schiner is the most economically important genus of fruit flies in Mexico (Aluja et al. 1987). These fruit flies are currently detected and surveyed with McPhail traps, although problems with efficiency and variability of capture have been described (Liedo 1983, Aluja et al. 1989). A few studies of the behavior of fruit flies employing McPhail traps have been reported (Prokopy & Economopoulos 1975, Aluja et al. 1989). McPhail (1937) studied the adult activity of Anastrepha ludens (Loew) in mango trees using McPhail traps This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.ufl.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2

Scientific Notes 291 with fermenting sugar as bait. The current work reports the capture of Anastrepha obliqua (Macquart) and A. ludens in relation to time of day using McPhail traps baited with proteinaceous attractant. This information may be useful in understanding the dynamics of fruit fly trapping. The study was carried out in the Mazapa de Madero Valley, Chiapas, Mexico, from 27 to 29 May and 12 to 16 June, 1986, with wild Anastrepha flies, using the same host trees in both periods. This work was conducted before a medfly program began to release parasitoids for biological control of Anastrepha fruit flies. The tests were done during the period when trap captures were normally at their peak (Aluja et al. 1989). Previous studies in Mazapa de Madero Valley have been described by Aluja et al. (1987) and Malo (1992). Traps were baited with a mixture of torula yeast and borax in a ratio of 4:5 (21 g) dissolved in 300 ml of water (Lopez et al. 1971). Twelve McPhail traps were hung at a height of 7 m in mango trees, Mangifera indica L. variety “criollo”, also known as “mango de coche”. The criteria for selection of trees in which to hang the traps were mostly concerned with factors such as ease of access and the presence of mature fruits (approximately 80% on the tree). The distance between traps was a minimum of 100 m. Traps were hung on the trees at 0600 hours and serviced every two hours, ending at 1800 hours. Traps were left hanging in trees throughout the night to determine the possible capture during this time of day. The contents of each trap were sieved, and the captured insects were rinsed with clean water and placed in a 50 ml vial filled with 70% ethanol; the traps were also rinsed and re-baited. The amount of bait used for the traps was kept constant during each 3 to 5 day-long observation period. Captured flies were counted and identified to species and sex using Steyskal's (1977) key. Temperatures and relative humidities were recorded with a hygrothermograph placed in the shade of a mango tree local- ized in the center of the valley. Wind velocity was measured every two hours with an anemometer placed in the canopy of the same mango tree (10 m height approximately). A total of 1,929 flies of the genus Anastrepha was captured, of which 56.8% were A. obliqua, 39.8% A. ludens and 3.4% other species such as A. serpentina (Wiede- mann), A. distincta Greene, A. fraterculus (Wiedemann) and A. striata Schiner. Be- cause the catches of A. obliqua and A. ludens amounted to 96.6% of all flies caught, the analysis was made with these species only. In earlier studies reported by Aluja et al. (1992), A. ludens was the species most abundant, followed by A. obliqua and A. serpentina in Mazapa de Madero Valley. Malo (1992) found that the predominant species were A. ludens and A. obliqua. The findings obtained in this work confirmed that the latter two species were the most abundant. Almost all the mangoes growing in the Valley were “mango de coche”, a variety highly preferred by these species (Aluja & Liedo, 1986). Data in Table 1 show the mean number (± S.E.) of A. obliqua and A. ludens captured per trap per 2-hour period as a function of the time of day, mean temperature, relative humidity and wind velocity. For both species, the first flies were caught at 0800 hours; the 2-hour catch rate increased to a maximum peak between 1400 to 1600 hours, then declined from 1600 to 1800 hours. A. obliqua captures showed a significant difference in relation to time of day (F = 4.21; df = 6,49; P ≤ 0.01). Similar variation was observed in A. ludens (F = 6.76; df = 6,49; P ≤ 0.01). There were no flies trapped in the period between 1800 to 0600 hours. Variations in temperature and wind velocity throughout the trapping period showed a similar profile to that of capture of fruit flies in relation of time of day. The relative humidity was high in the morning, decreased in the mid-day, and increased again in the evening. The catches of A. ludens and A. obliqua were probably influenced by all these factors. McPhail trap catches have been shown to be affected by bi-

292 Florida Entomologist 77(2) June, 1994 otic factors, both of the pest and host origin, and non-biotic factors (McPhail 1937, Neuenschwander & Michelakis 1979, Kapatos & Fletcher 1982). Of the total number of Anastrepha fruit flies caught, 1,216 were females and 713 were males, a male:female ratio of 1:1.7. These results confirm previous reports indi- cating that McPhail traps baited with a proteinaceous feeding attractant capture more Anastrepha females than males (Lopez & Hernandez-Becerril 1967, Houston 1981, Malo 1992). For both species, the greatest captures of females occurred at 1600 hours, whereas for males it took place at 1400 hours. However, the number of males and females captured per trap did not differ significantly (Table 1). McPhail (1937) found similar diurnal patterns for males and females of A. ludens, although more males than females were caught in this case. These differences between sexes in number caught may be the result of different volatiles produced by sugar fermentation and protein baits (Malo 1992). Finally, in spite of the disadvantages of the McPhail traps in terms of fragility, bulkiness, and need of water (Liedo 1983), these traps can be used to capture a great diversity of Anastrepha fruit flies. From these results, it is clear that tests with McPhail traps can be performed between 1000 and 1800 hours, disregarding the noc- turnal period. Better knowledge of the biology and behavior of fruit flies, as well as their preferences to different baits used, should permit the design of an improved trap system for management of fruit flies. The authors would like to thank Jorge Valenzuela (Instituto de Ecologia, Xalapa, Mexico) for helpful discussions of this work and J. Hendrichs (IAEA, Vienna, Aus- tria), J. Sivinski (IABBBRL, Gainesville, Florida) and J. Cibrian-Tovar (Centro de Entomologia y Acarologia, Colegio de Postgraduados de Chapingo, Texcoco, Mexico) for helpful comments on the manuscript. Technical assistance was provided by Miguel Angel Guzman, Gerardo Hernandez-Rojas and Rufino Vargas.

SUMMARY

McPhail trap capture of Anastrepha fruit ﬂies in relation to time of day was studied. Six species were caught, with Anastrepha obliqua and Anastrepha ludens being the predominant species. The ﬁrst capture was obtained at 0800 hours and catches increased throughout the day up to a maximum between 1400 and 1600 hours. More females than males of both species were captured.

REFERENCES CITED

ALUJA, M., AND P. LIEDO. 1986. Future perspectives on integrated management of fruit flies in Mexico, pp. 12-48 in M. Mangel, J. R. Carey and R. Plant [eds.]. Pest Control: operation and systems analysis in fruit fly management. Proc. NATO advanced workshop. Springer Verlag. New York. ALUJA, M., M. CABRERA, E. RIOS, J. GUILLEN, H. CELEDONIO, J. HENDRICHS, AND P. LIEDO. 1987. A survey of the economically important fruit flies (Diptera: Te- phritidae) present in Chiapas and a few other fruit regions in Mexico. Florida Entomol. 70: 320-329. ALUJA, M., M. CABRERA, J. GUILLEN, H. CELEDONIO, AND F. AYORA. 1989. Behaviour of Anastrepha ludens, A. obliqua and A. serpentina (Diptera: Tephritidae) on a wild mango tree (Mangifera indica) harbouring tree McPhail traps. Insect Sci. Applic. 10: 309-318. ALUJA, M., J. GUILLEN, P. LIEDO, M. CABRERA, E. RIOS, G. DE LA ROSA, H. CELE- DONIO, AND D. MOTA. 1990. Fruit infesting tephritids (Diptera: Tephritidae) and associated parasitoids in Chiapas, Mexico. Entomophaga. 35: 39-48. HOUSTON, W. W. K. 1981. Fluctuations in number and the significance of the sex ratio of the Mexican fruit Anastrepha ludens caught in McPhail traps. Entomol. exp. & appl. 30: 140-150. Scientific Notes 293 IN

DAY 3

* m/sec OF

Wind Velocity Wind TIME

3 * RELATION

IN C R. h. %

3 ° LUDENS

0.15 24 79 0 0.36 27 69 0.43 0.30 29 64 0.78 0.29 28 65 0.56 0.12 25 70 0.43 0.00 * 0.01 21 82 0.03 ± ± ± ± ± ± ± NASTREPHA A

1 AND

0.10 0.70 0.20 1.35 0.25 1.28 0.26 0.80 0.12 0.40 0.00 0.00 0.00 0.02 ± ± ± ± ± ± ± Males Females T OBLIQUA

Anastrepha ludens 2 NASTREPHA 0.25abc 0.35 0.52ab 0.84 0.53a 1.08 0.52abc 0.68 0.23bc 0.35 O.OOc 0.00 0.01c 0.00 A ± ± ± ± ± ± ±

TRAP

PER

0.42 1.06 0.84 2.19 0.59 2.37 0.58 1.57 0.24 0.75 0.00 0.00 0.01 0.02 ± ± ± ± ± ± ± . 1 EXICO CAPTURED

, M FLIES

0.22 0.96 0.45 2.42 0.50 1.96 0.15 1.46 0.05 0.64 0.00 0.00 0.00 0.02 ± ± ± ± ± ± ± HIAPAS OF Males Females Total , C S.E.) ± ( Anastrepha obliqua ADERO M 2 DE 1.24a 1.22 1.05a 1.41 0.00b 0.00 0.01b 0.00 0.64ab 0.46 0.71ab 0.49 0.28ab 0.18

± ± ± ± ± ± ± NUMBER

EAN AZAPA M Student t test unpaired with two tails, (p > 0.05) showed no signiﬁcant differences between males and females. ANOVA. test with Means in the same column followed by letter are not signiﬁcantly different at 5% level based on Tukey's These values were not obtained. 1 2 3 Day Total 18:00 1.43 16:00 3.65 14:00 3.38 12:00 1.96 10:00 0.83 06:00 0.00 Time of Time 08:00 0.02 TABLE 1. M 294 Florida Entomologist 77(2) June, 1994

KAPATOS, E., AND B. FLETCHER. 1982. Development of a pest management system for Dacus oleae in Corfu by utilizing ecological criteria. Proc. Intern. Symp. of CEC/IOBC on fruit flies of economic importance, Athens, Greece, Nov. 16-19. 593-602. LIEDO, P. 1983. Mexican fruit fly Anastrepha ludens (Loew): response to visual stimuli in the presence of pheromonal compounds. M. Sc. Thesis. University of Southampton, U.K. 69 pp. LOPEZ, D. F., AND O. HERNANDEZ-BECERRIL. 1967. Sodium borate inhibits decomposition of two protein hydrolysates attractive to Mexican fruit fly. J. Econ. En- tomol. 60: 136-140. LOPEZ, D. F., L. F. STEINER, AND F. R. HOLBOOK. 1971. A new yeast hydrolysate-borax bait for trapping the caribbean fruit fly. J. Econ. Entomol. 64:1541-1543. MALO, E. A. 1992. The effect of bait decomposition on the capture of Anastrepha fruit flies. Florida Entomol. 75: 272-274. MCPHAIL, M. 1937. Relation of time of day, temperature, and evaporation to attractiveness of fermenting sugar solution to Mexican fruit fly. J. Econ. Entomol. 30: 793-799. NEUENSCHWANDER, P., AND S. MICHELAKIS. 1979. McPhail trap captures of Dacus oleae (Gmel.) (Diptera: Tephritidae) in comparison to the fly density and population composition as assessed by sondage technique in Crete, Greece. Bull. Soc. Entomol. Suisse. 52: 343-357. PROKOPY, R. J., AND A. P. ECONOMOPOULOS. 1975. Attraction of laboratory-cultured and wild Dacus oleae flies to sticky-coated McPhail traps of different colors and odors. Environ. Entomol. 4: 187-192. STEYSKAL, G. C. 1977. Pictorial key to species of the genus Anastrepha (Diptera: Te- phritidae). Entomol. Soc. Washington. 35 pp.

Book Reviews 295

BOOK REVIEWS

COKENDOLPHER, JAMES C. AND VINCENT F. LEE. 1993. Catalogue of the Cyphopalpa- tores and Bibliography of the Harvestmen (Arachnida, Opiliones) of Greenland, Can- ada, U.S.A., and Mexico. Vintage Press, Lubbock, Texas. Distributed by The Wishing Well, 1200 Clover Drive, Burkburnett, TX 76354. iii + 82 p. Paperback, 17 X 24 cm, $9.50 includes shipping and handling; also available on 3.5" or 5.25" computer disk, MacIntosh or IBM, $20.00 includes shipping and handling. A brochure accompanied the review copy of this publication which contained an abbreviated form of the introduction, and since it effectively outlined the contents, it is reproduced below. “Harvestmen or daddy-long-legs are found above and below the ground in virtually every terrestrial habitat. They occur well past the Arctic Circle as far north as 73o in Canada. Even so, these often conspicuous and common animals are generally little studied or understood. Like most diverse groups of animals, the literature on Opiliones can be overwhelming. There are currently about 1400 publications (in most major languages of the world) dealing with the opilions of North America. The goal of this publication is to provide as complete as possible a listing of all of those ar- ticles (including newspapers, theses, dissertations, and government reports) on both fossil and recent species. All topics are covered including folklore. The literature surveys cover up to the end of 1992. Furthermore, a catalogue and geographical checklist are also provided to the 225 species and 50 genera of Cyphopalpatores (families Caddidae, Ceratolasmatidae, Nemastomatidae, Nemastomoididae, Phalangiidae, Protolophidae, Sabaconidae, Sclerosomatidae, Sironidae, Trogulidae) harvestmen from North America.” I have to agree that this publication is all it claims to be. It is arranged in four parts: a list of early names along with their current synonyms; a catalogue of all taxa; a checklist by state, province, and territory; and the bibliography. While the ad- vantages of having this information compiled together seem self-evident, one aspect which is often overlooked in similar compilations is the set of geographic checklists. Now anyone wanting to work on this poorly studied group will know exactly which species have been found in a particular area. The bibliography is exceptionally thor- ough including, as it does, a number of poorly accessible works. The authors point out that a bibliography of the West Indian harvestmen fauna was published recently by Cokendolpher & Camilo-Rivera (1989). That publication along with this one cover the bibliography of the entire opilion fauna of North Amer- ica. This catalog, along with that of Crawford (1992), covers the entire opilion no- menclature of North America. This privately published catalogue is remarkably free of errors, especially considering the difficulty of proofing the large number of printed scientific names within. It has good print quality, although perhaps not quite as good as the better scientific journals. The price ($9.50 for the book, $20.00 for computer disk) is a steal. If you have any interest at all in arachnids or biogeography, get both.

LITERATURE CITED COKENDOLPHER, J. C., AND G. C. CAMILO-RIVERA. 1989. Annotated bibliography to the harvestmen of the West Indies (Arachnida: Opiliones). Occas. Pap. Florida State Coll. Arthropods 5: 1-20. CRAWFORD, R. 1992. Catalogue of the genera and type species of the harvestman su- perfamily Phalangioidea (Arachnida). Burke Museum Contrib. Anthropol. Natur. Hist. 8: 1-60. G. B. EDWARDS Division of Plant Industry Florida State Collection of Arthropods P. O. Box 147100, Gainesville, FL 32614-7100

296 Florida Entomologist 77(2) June, 1994

WISE, D. H. 1993. Spiders in ecological webs. Cambridge University Press [Cam- bridge Studies in Ecology], Cambridge, Great Britain. Available from the Press Syn- dicate of the University of Cambridge, 40 West 20th Street, New York, NY 10011- 4211. ISBN 0-521-32547-1. xiii + 328 p. Hardback with dust cover, 15.5 X 23.5 cm. $79.95.

There has long been a need for a synthesis and evaluation of the work that has been done on spider ecology. This book makes a good start on filling that need. The first of nine chapters begins with a short introduction to spiders and brief general- ized outlines of the life histories of some of the more common families. A short account of spider mythology and the derivation of the word “spider” is also included. Lastly, there are discussions of spiders as model terrestrial predators and the importance of field experimentation, which is the primary focus of this book. The following chapters deal with food limitations (many spiders may be frequently hungry); the evidence for and against interspecific competition; lack of support for the classical interspecific competitionist paradigm in most field studies; how spiders avoid competition (little evidence for intraspecific competition); the impact of spiders on insect populations (including in agroecosystems); effects of environmental structure on spider populations; spiders as part of the larger community and effects of spider guilds on prey populations; and finally, a helpful discussion on experimental design and common errors (some of which the author reports he has been previously guilty of himself!), along with suggestions for future research. Useful synopses occur at the end of each chapter. The author points out that most of the ecological work on spiders has been done on those species which build webs because these species are more easily manipulated than wandering spiders. Unfortunately, the level of knowledge on wandering spiders is illustrated on p. 10, where the author states that salticids cannot capture prey in the dark. This primarily diurnal, visually-oriented family can capture prey in the dark by tactile means. Despite this and a few other minor exceptions, the book is remarkably free of errors. The quality of the production is high. This is reflected in the price of the book, which is in the range of comparable volumes, in other words, is fairly expensive. The bibliography is extensive, although due to the focus of the book on field experiments, not exhaustive. Considerable purely observational field work has not been included. The book is written in a metaphorical style which seems to be an attempt to broaden the potential audience for the book; however, the breadth and depth of the research presented and analyzed here will require a fair background in biology to be fully appreciated. Nevertheless, I would heartily recommend this book as the place to start for anyone interested in doing serious work in the field of spider ecology. It would be worth the time of an ecologist working on any taxon to read the last chapter before starting a field experiment.

G. B. Edwards Division of Plant Industry Florida State Collection of Arthropods P. O. Box 147100 Gainesville, FL 32614-7100

Book Reviews 297

S. R. LEATHER, K. F. A. WALTERS AND J. S. BALE. 1993. The ecology of insect overwintering. Cambridge University Press; Cambridge, England. Hardback, x + 255 p. ISBN 0-521-41758-9. $59.95.

As I write this review, the air temperature this December morning hovers about 5oC, a chilly reminder that even southern Florida feels the impact of winter. Lacking insulating clothes, our local insect fauna must practice some of the behavioral or metabolic adaptations that are the topics of the present book by three British inves- tigators. This attractive volume stresses physiological mechanisms and draws from a somewhat different journal literature compared with its closest facsimile, Sea- sonal adaptations of insects by Tauber et al. (1986), which presents a more evolutionary approach to overwintering. Following a brief Introduction, Chapter 2 of Leather et al. considers “The overwintering locale - suitability and selection”, a unique overview of microhabitats and mi- croclimates sought by insects for protection from hibernal conditions. This chapter culls a diverse literature, including many relevant citations from meteorology and even one from a speleology journal, that bear on overwintering sites. The third chapter “Stimuli controlling diapause and overwintering” documents an impressive variety of entomological responses to seasonal environmental cues. This section describes how insects use daylength, temperature, and, to a lesser extent, biotic stimuli in mul- tifarious ways to regulate diapause initiation, maintenance, or termination. Chapter 4 on “Insect cold-hardiness” is the longest (76 pages) and, arguably, most specialized portion of the book, focusing on mechanisms of cryoprotection. In spite of stated goals to merge ecological and physiological views of cold-hardiness, this chapter dwells so intensively on physiological and biochemical specializations that the connecting threads to insect ecology are barely visible. Nevertheless, this section provides excellent coverage of insect cryobiology, even borrowing from fish (anti- freeze proteins were first discovered from cold-water species) and plant physiology to describe cryo-protective mechanisms. “Costs and benefits of overwintering” is the ti- tle of Chapter 5 which examines an eclectic mix of themes such as familial or latitu- dinal specificity of overwintering stages, bet-hedging strategies, and the physical, metabolic, and reproductive costs of overwintering. The final chapter “Prediction and control” describes circumstances (aphid and moth pests receive most coverage) where counts of pre-winter abundance have been used to forecast the probability and intensity of a post-diapause outbreak. Examples of host-plant manipulations and cultural practices, such as tillage, are recommended as alternatives to direct control to forestall an outbreak. Although the authors advocate a detailed understanding of the overwintering process to facilitate control, simple life table information would suffice to predict the emergence levels of pests based upon pre-winter abundances. An appealing feature of this book is its use of case-study methods to discuss overwintering paradigms in depth. The many European examples, drawing from publications in Scandinavian tongues as well as Russian, French and German, should provide some fresh insights to North American readers. Graphs and figures, some original and others reproduced from journals, are distributed liberally through the text to elucidate concepts or highlight data. Some graphs are, unfortunately, difficult to decipher or provided with insufficient documentation. Other irritations include missing or inaccurate citations and some ponderous sentence structures. Yet, these are minor quibbles about a valuable book that fills a vacant niche on insect cryobiology and should stimulate entomologists to consider the pure and applied ramifications of overwintering strategies.

L. P. Lounibos Florida Medical Entomology Lab University of Florida 200 9th St. SE, Vero Beach, FL 32962 This article is from Florida Entomologist Online, Vol. 77, No. 2 (1994). FEO is available from the Florida Center for Library Automation gopher (sally.fcla.uﬂ.edu) and is identical to Florida Entomologist (An International Journal for the Americas). FEO is prepared by E. O. Painter Printing Co., P.O. Box 877, DeLeon Springs, FL. 32130. This document was created with FrameMaker 4.0.2